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Sample records for cells produce nerve

  1. Bone marrow cells produce nerve growth factor and promote angiogenesis around transplanted islets

    Institute of Scientific and Technical Information of China (English)

    Naoaki; Sakata; Nathaniel; K; Chan; John; Chrisler; Andre; Obenaus; Eba; Hathout

    2010-01-01

    AIM:To clarify the mechanism by which bone marrow cells promote angiogenesis around transplanted islets.METHODS: Streptozotocin induced diabetic BALB/ c mice were transplanted syngeneically under the kidney capsule with the following: (1) 200 islets (islet group: n=12), (2) 1-5×106 bone marrow cells (bone marrow group: n=11), (3) 200 islets and 1-5×106 bone marrow cells (islet + bone marrow group: n= 13), or (4) no cells (sham group:n=5). All mice were evaluated for blood glucose, serum insulin, serum nerve...

  2. A Cell Line Producing Recombinant Nerve Growth Factor Evokes Growth Responses in Intrinsic and Grafted Central Cholinergic Neurons

    Science.gov (United States)

    Ernfors, Patrik; Ebendal, Ted; Olson, Lars; Mouton, Peter; Stromberg, Ingrid; Persson, Hakan

    1989-06-01

    The rat β nerve growth factor (NGF) gene was inserted into a mammalian expression vector and cotransfected with a plasmid conferring resistance to neomycin into mouse 3T3 fibroblasts. From this transfection a stable cell line was selected that contains several hundred copies of the rat NGF gene and produces excess levels of recombinant NGF. Such genetically modified cells were implanted into the rat brain as a probe for in vivo effects of NGF on central nervous system neurons. In a model of the cortical cholinergic deficits in Alzheimer disease, we demonstrate a marked increase in the survival of, and fiber outgrowth from, grafts of fetal basal forebrain cholinergic neurons, as well as stimulation of fiber formation by intact adult intrinsic cholinergic circuits in the cerebral cortex. Adult cholinergic interneurons in intact striatum also sprout vigorously toward implanted fibroblasts. Our results suggest that this model has implications for future treatment of neurodegenerative diseases.

  3. Neuronal differentiation of PC12 cells induced by sciatic nerve and optic nerve conditioned medium

    Institute of Scientific and Technical Information of China (English)

    DU Chan; YANG De-mei; ZHANG Pei-xun; DENG Lei; JIANG Bao-guo

    2010-01-01

    Background Previous work has shown that optic nerve and sciatic nerve conditional medium had neurotrophic activity on neurons. In order to find if the optic nerve conditioned media (CM) had a similar activity to make PC12 cells differentiate as sciatic nerve CM did, we explored the neurotrophic activity in optic nerve CM in the same in vitro system and compared the neurotrophin expression levels in optic and sciatic nerves under both conditions.Methods PC12 cells were used to examine the effects of neurotrophins secreted by the sciatic nerve and optic nerve. RT-PCR and real-time QPCR showed that the sciatic nerve and optic nerve produced a range of neurotrophins including nerve growth factor (NGF), brain derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3).Results The effects of sciatic nerve and optic nerve CM on neurite outgrowth were tested against a range of neurotrophins, and they had different neuritogenic activities. Only NGF and sciatic nerve CM had obvious neuritogenic activities, although the concentration of NGF in the sciatic nerve CM was very low.Conclusions Our experiment showed that sciatic nerve CM had a higher neurotrophic activity on PC12 cells than optic nerve CM. These results suggested that peripheral nervous system (PNS) and central nervous system (CNS) had different expression levels of neurotrophin, which may in part explain the lack of ability to regenerate the CNS.

  4. Fibroblasts that reside in mouse and frog injured peripheral nerves produce apolipoproteins.

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    Saada, A; Dunaevsky-Hutt, A; Aamar, A; Reichert, F; Rotshenker, S

    1995-05-01

    Apolipoprotein synthesis and secretion is upregulated in wallerian degenerating peripheral nerves. A commonly expressed view has been that macrophages are solely responsible for their production. In the present study we provide evidence that (1) nerve-derived fibroblasts contribute to apolipoprotein production, (2) apolipoprotein production is confined to regions where myelin destruction and phagocytosis occur, and (3) some experimental procedures are detrimental for the production of apolipoproteins. Apolipoprotein production was studied in C57BL/6/NHSD (N) and C57/BL/6-WLD/OLA/NHSD (W) mice that display, respectively, rapid and slow progression of wallerian degeneration. In N nerves, apolipoprotein E (apo-E) is produced during in vitro and in vivo degeneration, and in vivo after freeze damage. In W nerves, apo-E is produced at the injury region where degeneration occurs but not farther distally where degeneration fails to develop. Apo-E is also produced in W nerves during in vitro degeneration and in vivo after freeze damage. In culture, N and W mice nerve-derived fibroblasts, but neither macrophages nor Schwann cells produced apo-E. Two apolipoproteins are produced in in vivo wallerian degenerating and freeze-damaged frog nerves, i.e., apo-39 and apo-29. Only apo-39 is produced in in vitro degenerating nerves. Neither apo-39 nor apo-29 is produced during in vivo degeneration in diffusion chambers. In culture, apo-39 is produced by nerve-derived fibroblasts and macrophages but not by Schwann cells.

  5. Adipose derived stem cells and nerve regeneration

    Institute of Scientific and Technical Information of China (English)

    Alessandro Faroni; Richard JP Smith; Adam J Reid

    2014-01-01

    Injuries to peripheral nerves are common and cause life-changing problems for patients along-side high social and health care costs for society. Current clinical treatment of peripheral nerve injuries predominantly relies on sacriifcing a section of nerve from elsewhere in the body to pro-vide a graft at the injury site. Much work has been done to develop a bioengineered nerve graft, precluding sacriifce of a functional nerve. Stem cells are prime candidates as accelerators of re-generation in these nerve grafts. This review examines the potential of adipose-derived stem cells to improve nerve repair assisted by bioengineered nerve grafts.

  6. On the use of upper extremity proximal nerve action potentials in the localization of focal nerve lesions producing axonotmesis.

    Science.gov (United States)

    White, J C

    1997-09-01

    Ulnar, median, and radial proximal nerve action potentials (PNAPs) were recorded from the axilla and supraclavicularly, with stimulation of the nerves at the elbow or the radial groove, in 30 control subjects for each nerve. In addition to routine nerve conduction studies, wrist to elbow median nerve action potentials were recorded proximal to the lesion in 76 patients with carpal tunnel syndrome of varying degrees of severity to determine the effect that the distal lesion might have on more proximal nerve conduction. Utilizing this information, PNAPs, standard nerve conduction studies, and needle electrode examinations were carried out in patients with focal elbow area nerve or brachial plexus lesions producing axonotmesis. PNAPs confirmed the site of the lesions producing axonotmesis when localization was possible with standard nerve conduction and/or needle electrode studies and were the sole means by which localization of the lesions producing only sensory axonotmesis was accomplished.

  7. Acellular allogeneic nerve grafting combined with bone marrow mesenchymal stem cell transplantation for the repair of long-segment sciatic nerve defects: biomechanics and validation of mathematical models

    Directory of Open Access Journals (Sweden)

    Ya-jun Li

    2016-01-01

    Full Text Available We hypothesized that a chemically extracted acellular allogeneic nerve graft used in combination with bone marrow mesenchymal stem cell transplantation would be an effective treatment for long-segment sciatic nerve defects. To test this, we established rabbit models of 30 mm sciatic nerve defects, and treated them using either an autograft or a chemically decellularized allogeneic nerve graft with or without simultaneous transplantation of bone marrow mesenchymal stem cells. We compared the tensile properties, electrophysiological function and morphology of the damaged nerve in each group. Sciatic nerves repaired by the allogeneic nerve graft combined with stem cell transplantation showed better recovery than those repaired by the acellular allogeneic nerve graft alone, and produced similar results to those observed with the autograft. These findings confirm that a chemically extracted acellular allogeneic nerve graft combined with transplantation of bone marrow mesenchymal stem cells is an effective method of repairing long-segment sciatic nerve defects.

  8. GRP-producing nerves control antral somatostatin and gastrin secretion in pigs

    DEFF Research Database (Denmark)

    Holst, J J; Orskov, C; Poulsen, Steen Seier

    1987-01-01

    of isolated perfused pig antrum with intact vagus nerve supply. Electrical stimulation of the vagus nerves at 4 Hz increased the antral release of GRP up to 10-fold and increased SS output 2- to 3-fold. Atropine at 10(-6) M had no effect on these responses. Intra-arterial GRP increased SS secretion...... the effects of vagus stimulation on gastrin and somatostatin output. Gastrin in concentrations up to 10(-7) M was without effect on SS secretion. We conclude that electrical stimulation of the vagus nerves increases antral SS gastrin secretion and that GRP is a likely transmitter.......By immunohistochemistry, nerve fibers containing gastrin-releasing polypeptide (GRP)-like immunoreactivity were identified close to the somatostatin (SS)-producing cells of the gastric antral mucosa. We, therefore, studied the possible role of GRP in the control of antral SS secretion by use...

  9. Locally Produced BDNF Promotes Sclerotic Change in Alveolar Bone after Nerve Injury

    Science.gov (United States)

    Ida-Yonemochi, Hiroko; Yamada, Yurie; Yoshikawa, Hiroyuki

    2017-01-01

    Brain-derived neurotrophic factor (BDNF), which is released due to nerve injury, is known to promote the natural healing of injured nerves. It is often observed that damage of mandibular canal induces local sclerotic changes in alveolar bone. We reported that peripheral nerve injury promotes the local production of BDNF; therefore, it was possible to hypothesize that peripheral nerve injury affects sclerotic changes in the alveolar bone. This study aimed to evaluate the effect of BDNF on osteogenesis using in vitro osteoblast-lineage cell culture and an in vivo rat osteotomy model. MC3T3-E1 cells were cultured with BDNF and were examined for cell proliferative activity, chemotaxis and mRNA expression levels of osteoblast differentiation markers. For in vivo study, inferior alveolar nerve (IAN) injury experiments and mandibular cortical osteotomy were performed using a rat model. In the osteotomy model, exogenous BDNF was applied to bone surfaces after corticotomy of the mandible, and we morphologically analyzed the new bone formation. As a result, mRNA expression of osteoblast differentiation marker, osteocalcin, was significantly increased by BDNF, although cell proliferation and migration were not affected. In the in vivo study, osteopontin-positive new bone formation was significantly accelerated in the BDNF-grafted groups, and active bone remodeling, involving trkB-positive osteoblasts and osteocytes, continued after 28 days. In conclusion, BDNF stimulated the differentiation of MC3T3-E1 cells and it promoted new bone formation and maturation. These results suggested that local BDNF produced by peripheral nerve injury contributes to accelerating sclerotic changes in the alveolar bone. PMID:28072837

  10. Research Status of Astragali Radix on Nerve Cells and Nerve System Diseases

    Institute of Scientific and Technical Information of China (English)

    LUAN Zeng-qiang; ZHAO Ping-li; CAO Wen-fu

    2012-01-01

    Astragali Radix has a wide application in the nerve system diseases because of its obvious nerve cell protection and recovery effects.Astragali Radix has good clinical effects both in acute and chronic cerebrovascular diseases and neurological degenerative diseases.This paper reviews the experimental and clinical research status of Astragali Radix on nerve system and nerve system diseases,which may promote its experimental research and clinical application.

  11. Sericin protects against diabetes-induced injuries in sciatic nerve and related nerve cells

    Institute of Scientific and Technical Information of China (English)

    Chengjun Song; Zhenjun Yang; Meirong Zhong; Zhihong Chen

    2013-01-01

    Sericin from discarded silkworm cocoons of silk reeling has been used in different fields, such as cosmetology, skin care, nutrition, and oncology. The present study established a rat model of type 2 diabetes by consecutive intraperitoneal injections of low-dose (25 mg/kg) streptozotocin. After intragastrical perfusion of sericin for 35 days, blood glucose levels significantly declined, and the expression of neurofilament protein in the sciatic nerve and nerve growth factor in L4–6 spinal ganglion and anterior horn cells significantly increased. However, the expression of neuropeptide Y in spinal ganglion and anterior horn cells significantly decreased in model rats. These findings indicate that sericin protected the sciatic nerve and related nerve cells against injury in a rat type 2 diabetic model by upregulating the expression of neurofilament protein in the sciatic nerve and nerve growth factor in spinal ganglion and anterior horn cells, and downregulating the expression of neuropeptide Y in spinal ganglion and anterior horn cells.

  12. Acellular allogeneic nerve grafting combined with bone marrow mesenchymal stem cell transplantation for the repair of long-segment sciatic nerve defects:biomechanics and validation of mathematical models

    Institute of Scientific and Technical Information of China (English)

    Ya-jun Li; Bao-lin Zhao; Hao-ze Lv; Zhi-gang Qin; Min Luo

    2016-01-01

    We hypothesized that a chemically extracted acellular allogeneic nerve graft used in combination with bone marrow mesenchymal stem cell transplantation would be an effective treatment for long-segment sciatic nerve defects. To test this, we established rabbit models of 30 mm sciatic nerve defects, and treated them using either an autograft or a chemically decellularized allogeneic nerve graft with or without simultaneous transplantation of bone marrow mesenchymal stem cells. We compared the tensile properties, electrophysiological function and morphology of the damaged nerve in each group. Sciatic nerves repaired by the allogeneic nerve graft combined with stem cell trans-plantation showed better recovery than those repaired by the acellular allogeneic nerve graft alone, and produced similar results to those observed with the autograft. These ifndings conifrm that a chemically extracted acellular allogeneic nerve graft combined with transplanta-tion of bone marrow mesenchymal stem cells is an effective method of repairing long-segment sciatic nerve defects.

  13. Mechanism of histopathological changes of nerve cells experimentally induced by chronic alcohol poisoning.

    Science.gov (United States)

    Miyakawa, T; Sumiyoshi, S; Deshimaru, M; Hattori, E; Shikai, I

    1977-01-01

    Experimental alcoholism was produced in rats by supplying them with 15% ethanol as the only source of liquid for a whole year. Histopathological examination revealed that Purkinje cells and granule cells in the cerebellum mainly showed such changes as decrease of ER, ribosomes and severe atrophy of the nerve cells. It might be speculated that these changes were caused by the disturbance of protein synthesis in the nerve cells induced by chronic alcohol effect.

  14. Nerve Growth Factor is Primarily Produced by GABAergic Neurons of the Rat Neocortex

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    Jeremy eBiane

    2014-08-01

    Full Text Available Within the cortex, nerve growth factor (NGF mediates the innervation of cholinergic neurons during development, maintains cholinergic corticopetal projections during adulthood and modulates cholinergic function through phenotypic control of the cholinergic gene locus. Recent studies suggest NGF may also play an important role in cortical plasticity in adulthood. Previously, NGF-producing cells have been shown to colocalize with GABAergic cell markers within the hippocampus, striatum, and basal forebrain. Classification of cells producing NGF in the cortex is lacking, however, and cholinergic corticopetal projections have been shown to innervate both pyramidal and GABAergic neurons in the cortex. In order to clarify potential trophic interactions between cortical neurons and cholinergic projections, we used double-fluorescent immunohistochemistry to classify NGF-expressing cells in several cortical regions, including the prefrontal cortex, primary motor cortex, parietal cortex and temporal cortex. Our results show that NGF colocalizes extensively with GABAergic cell markers in all cortical regions examined, with >91% of NGF-labeled cells coexpressing GAD65/67. Conversely, NGF-labeled cells exhibit very little co-localization with the excitatory cell marker CaMKIIα (less than 5% of cells expressing NGF. NGF expression was present in 56% of GAD-labeled cells, suggesting that production is confined to a specific subset of GABAergic neurons. These findings demonstrate that GABAergic cells are the primary source of NGF production in the cortex, and likely support the maintenance and function of basal forebrain cholinergic projections in adulthood.

  15. Nerve growth factor interactions with mast cells.

    Science.gov (United States)

    Kritas, S K; Caraffa, A; Antinolfi, P; Saggini, A; Pantalone, A; Rosati, M; Tei, M; Speziali, A; Saggini, R; Pandolfi, F; Cerulli, G; Conti, P

    2014-01-01

    Neuropeptides are involved in neurogenic inflammation where there is vasodilation and plasma protein extravasion in response to this stimulus. Nerve growth factor (NGF), identified by Rita Levi Montalcini, is a neurotrophin family compound which is important for survival of nociceptive neurons during their development. Therefore, NGF is an important neuropeptide which mediates the development and functions of the central and peripheral nervous system. It also exerts its proinflammatory action, not only on mast cells but also in B and T cells, neutrophils and eosinophils. Human mast cells can be activated by neuropeptides to release potent mediators of inflammation, and they are found throughout the body, especially near blood vessels, epithelial tissue and nerves. Mast cells generate and release NGF after degranulation and they are involved in iperalgesia, neuroimmune interactions and tissue inflammation. NGF is also a potent degranulation factor for mast cells in vitro and in vivo, promoting differentiation and maturation of these cells and their precursor, acting as a co-factor with interleukin-3. In conclusion, these studies are focused on cross-talk between neuropeptide NGF and inflammatory mast cells.

  16. Genetic instability in nerve sheath cell tumors

    DEFF Research Database (Denmark)

    Rogatto, Silvia Regina; Casartelli, Cacilda; Rainho, Claudia Aparecida

    1995-01-01

    After in vitro culture, we analyzed cytogenetically four acoustic nerve neurinomas, one intraspinal neurinoma and one neurofibroma obtainedfrom unrelated patients. Monosomy of chromosomes 22 and 16 was an abnormality common to all cases, followed in frequency by loss of chromosomes 18 (three cases......) and chromosomes 8, 17 and 19 (two cases). Trisomy of chromosome 20 was also detected in two cases. Structural rearrangements were detected at low frequencies, with del(10)(p12) being present in two cases. In addition, we observed cell subpopulations showing a certain degree of genetic instability, reflected...

  17. Expression changes of nerve cell adhesion molecules L1 and semaphorin 3A after peripheral nerve injury

    Directory of Open Access Journals (Sweden)

    Qian-ru He

    2016-01-01

    Full Text Available The expression of nerve cell adhesion molecule L1 in the neuronal growth cone of the central nervous system is strongly associated with the direction of growth of the axon, but its role in the regeneration of the peripheral nerve is still unknown. This study explored the problem in a femoral nerve section model in rats. L1 and semaphorin 3A mRNA and protein expressions were measured over the 4-week recovery period. Quantitative polymerase chain reaction showed that nerve cell adhesion molecule L1 expression was higher in the sensory nerves than in motor nerves at 2 weeks after injury, but vice versa for the expression of semaphorin 3A. Western blot assay results demonstrated that nerve cell adhesion molecule L1 expression was higher in motor nerves than in the sensory nerves at the proximal end after injury, but its expression was greater in the sensory nerves at 2 weeks. Semaphorin 3A expression was higher in the motor nerves than in the sensory nerves at 3 days and 1 week after injury. Nerve cell adhesion molecule L1 and semaphorin 3A expressions at the distal end were higher in the motor nerves than in the sensory nerves at 3 days, 1 and 2 weeks. Immunohistochemical staining results showed that nerve cell adhesion molecule L1 expression at the proximal end was greater in the sensory nerves than in the motor nerves; semaphorin 3A expression was higher in the motor nerves than in the sensory nerves at 2 weeks after injury. Taken together, these results indicated that nerve cell adhesion molecules L1 and semaphorin 3A exhibited different expression patterns at the proximal and distal ends of sensory and motor nerves, and play a coordinating role in neural chemotaxis regeneration.

  18. Ulnar nerve injuries of the hand producing intrinsic muscle denervation of magnetic resonance imaging

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    Barberie, J.E.; Connell, D.G.; Munk, P.L.; Janzen, D.L. [Vancouver General Hospital and University of British Columbia, Vancouver, British Columbia, (Canada). Department of Radiology

    1999-08-01

    Muscle and nerve injuries in the hand may be difficult to detect and diagnose clinically. Two cases are reported in which magnetic resonance imaging showed ulnar nerve injury and intrinsic hand muscle denervation. The clinical, anatomical and radiological features of injury to the deep motor branch of the ulnar nerve and associated muscle denervation are discussed and illustrated. Compression of the deep motor branch of the ulnar nerve is a rare cause of hand dysfunction. This condition produces a clinical syndrome characterized by weakness of the muscles innervated by the deep branch of the ulnar nerve, with normal sensation in the hand. Denervation of the intrinsic hand muscles is variable, depending on the site and severity of the nerve injury. The anatomy of the ulnar nerve is complex and ulnar nerve compression syndrome is difficult to detect and localize clinically. Multiple causes exist, including ganglions; repeated trauma, typically occupationally related; calcification adjacent to the pisiform and the pisotriquetral joint; anomalous muscle bellies; fractures or dislocations affecting the ulnar side of the wrist; vascular anomalies of the ulnar artery; and oedema of the hand, requiring differing surgical treatments. We report two patients who presented with ulnar nerve compression syndrome secondary to post-traumatic neuromas as a result of blunt trauma to the palm. Copyright (1999) Blackwell Science Pty Ltd 12 refs., 2 figs.

  19. Which form of collagen is suitable for nerve cell culture?*

    Institute of Scientific and Technical Information of China (English)

    Mohsen Fathi Najafi; Saber Zahri; Fatemeh Vahedi; Leila Esmaililian Toosi; Nazila Ariaee

    2013-01-01

    In this study, we investigated the effects of hydrolyzed and non-hydrolyzed col agen and two-dimensional and three-dimensional col agen matrices on cell survival, attachment and neurite outgrowth of primary cultured nerve cells using 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) colorimetric assay and inverted microscopy. Hydrolyzed col agen facilitated nerve cell survival and neurite outgrowth, but it had no obvious influences on cellattachment. In contrast, non-hydrolyzed two-dimensional collagen matrix had no obvious effects on neurite outgrowth. These findings suggest that hydrolyzed col agen is an ideal nerve cell culture media.

  20. Platelet-rich plasma-induced bone marrow mesenchymal stem cells versus autologous nerve grafting for sciatic nerve repair

    Institute of Scientific and Technical Information of China (English)

    Changsuo Xia; Yajuan Li; Wen Cao; Zhaohua Yu

    2010-01-01

    Autologous nerve grafting is the gold standard of peripheral nerve repair.We previously showed that autologous platelet-rich plasma(PRP)contains high concentrations of growth factors and can induce in vitro cultured bone marrow mesenchymal stem cells(BMSCs)to differentiate into Schwann cells.Here we used PRP-induced BMSCs combined with chemically extracted acellular nerves to repair sciatic nerve defects and compared the effect with autologous nerve grafting.The BMSCs and chemically extracted acellular nerve promoted target muscle wet weight restoration,motor nerve conduction velocity,and axonal and myelin sheath regeneration,with similar effectiveness to autologous nerve grafting.This finding suggests that PRP induced BMSCs can be used to repair peripheral nerve defects.

  1. Dynamic regulation of Schwann cell enhancers after peripheral nerve injury.

    Science.gov (United States)

    Hung, Holly A; Sun, Guannan; Keles, Sunduz; Svaren, John

    2015-03-13

    Myelination of the peripheral nervous system is required for axonal function and long term stability. After peripheral nerve injury, Schwann cells transition from axon myelination to a demyelinated state that supports neuronal survival and ultimately remyelination of axons. Reprogramming of gene expression patterns during development and injury responses is shaped by the actions of distal regulatory elements that integrate the actions of multiple transcription factors. We used ChIP-seq to measure changes in histone H3K27 acetylation, a mark of active enhancers, to identify enhancers in myelinating rat peripheral nerve and their dynamics after demyelinating nerve injury. Analysis of injury-induced enhancers identified enriched motifs for c-Jun, a transcription factor required for Schwann cells to support nerve regeneration. We identify a c-Jun-bound enhancer in the gene for Runx2, a transcription factor induced after nerve injury, and we show that Runx2 is required for activation of other induced genes. In contrast, enhancers that lose H3K27ac after nerve injury are enriched for binding sites of the Sox10 and early growth response 2 (Egr2/Krox20) transcription factors, which are critical determinants of Schwann cell differentiation. Egr2 expression is lost after nerve injury, and many Egr2-binding sites lose H3K27ac after nerve injury. However, the majority of Egr2-bound enhancers retain H3K27ac, indicating that other transcription factors maintain active enhancer status after nerve injury. The global epigenomic changes in H3K27ac deposition pinpoint dynamic changes in enhancers that mediate the effects of transcription factors that control Schwann cell myelination and peripheral nervous system responses to nerve injury.

  2. Histological observation on acellular nerve grafts co-cultured with Schwann cells for repairing defects of the sciatic nerve

    Institute of Scientific and Technical Information of China (English)

    Xiaohong Sun; Jiangyi Tian; Xiaojie Tong; Xu Zhang; Zheng He

    2006-01-01

    BACKGROUND: Animal experiments and clinical studies about tissue engineering method applied to repair nerve injury mainly focus on seeking ideal artificial nerve grafts, nerve conduit and seed cells. Autologous nerve, allogeneic nerve and xenogeneic nerve are used to bridge nerve defects, it is one of the methods to promote the repair of nerve injury by culturing and growing Schwann cells, which can secrete various neurotrophic factor activities, in the grafts.OBJECTIVE: To observe the effect of acellular nerve grafts co-cultured with Schwann cells in repairing defects of sciatic nerve.DESIGN: An observational comparative study.SETTING: Tissue Engineering Laboratory of China Medical University.MATERIALS: The experiment was carried out in the Tissue Engineering Laboratory of China Medical University between April 2004 and April 2005. Forty neonatal Sprague-Dawley rats of 5-8 days (either males or females) and 24 male Wistar rats of 180-220 g were provided by the experimental animal center of China Medical University.METHODS: ① Culture of Schwann cells: The bilateral sciatic nerves and branchial plexus were isolated from the 40 neonatal SD rats. The sciatic nerves were enzymatically digested with collagenase and dispase, isolatd, purified and cultured with the method of speed-difference adhersion, and identified with the SABC immunohistochemical method. ② Model establishment: In vitro Schwann cells were microinjected into 10-mm long acellular nerve grafts repairing a surgically created gap in the rat sciatic nerve.According to the different grafted methods, the animals were randomly divided into three groups: autografts (n=8), acellular nerve grafts (n=8), or acellular nerve grafts with Schwann cells (n=8). ③ The regenerated nerve fiber number and average diameter of myeline sheath after culture were statistically anlayzed.MAIN OUTCOME MEASURES: ① The regenerated nerve ultrastructure, total number and density of myelinated nerve fibers, and the thickness of

  3. Hydrogen sulfide is essential for Schwann cell responses to peripheral nerve injury.

    Science.gov (United States)

    Park, Byung Sun; Kim, Hyun-Wook; Rhyu, Im Joo; Park, Chan; Yeo, Seung Geun; Huh, Youngbuhm; Jeong, Na Young; Jung, Junyang

    2015-01-01

    Hydrogen sulfide (H2 S) functions as a physiological gas transmitter in both normal and pathophysiological cellular events. H2 S is produced from substances by three enzymes: cystathionine β-synthase (CBS), cystathionine γ-lyase (CSE), and 3-mercaptopyruvate sulfurtransferase (MST). In human tissues, these enzymes are involved in tissue-specific biochemical pathways for H2 S production. For example, CBS and cysteine aminotransferase/MST are present in the brain, but CSE is not. Thus, we examined the expression of H2 S production-related enzymes in peripheral nerves. Here, we found that CSE and MST/cysteine aminotransferase, but not CBS, were present in normal peripheral nerves. In addition, injured sciatic nerves in vivo up-regulated CSE in Schwann cells during Wallerian degeneration (WD); however, CSE was not up-regulated in peripheral axons. Using an ex vivo sciatic nerve explant culture, we found that the inhibition of H2 S production broadly prevented the process of nerve degeneration, including myelin fragmentation, axonal degradation, Schwann cell dedifferentiation, and Schwann cell proliferation in vitro and in vivo. Thus, these results indicate that H2 S signaling is essential for Schwann cell responses to peripheral nerve injury. Hydrogen sulfide (H2 S) functions as a physiological gas transmitter in both normal and pathophysiological cellular events. H2 S is produced from cystathionine β-synthase (CBS), cystathionine γ-lyase (CSE), and 3-mercaptopyruvate sulfur transferase (MST). Here, we found that CSE and MST/CAT were present in normal peripheral nerves. Injured static nerves in vivo up-regulated CSE in Schwann cells during Wallerian degeneration, but CSE was not up-regulated in peripheral axons.

  4. Peripheral nerve extract effects on mesenchymal cells.

    OpenAIRE

    Dietz, F. R.; Mukhopadhyay, B.; Becker, G.; Daniels, K.; Solursh, M

    1996-01-01

    Several common congenital limb disorders are characterized by normal tissue differentiation but abnormal somatic growth. These include: idiopathic clubfoot, idiopathic leg length discrepancy, hemi-atrophy and hemi-hypertrophy. Both clinical and research studies have suggested that peripheral nerves may be important in regulating somatic growth of limb tissues. To investigate the hypothesis that peripheral nerves convey trophic substances to mesenchymal tissues that are involved in the regulat...

  5. Endocrine cells producing regulatory peptides.

    Science.gov (United States)

    Solcia, E; Usellini, L; Buffa, R; Rindi, G; Villani, L; Zampatti, C; Silini, E

    1987-07-15

    Recent data on the immunolocalization of regulatory peptides and related propeptide sequences in endocrine cells and tumors of the gastrointestinal tract, pancreas, lung, thyroid, pituitary (ACTH and opioids), adrenals and paraganglia have been revised and discussed. Gastrin, xenopsin, cholecystokinin (CCK), somatostatin, motilin, secretin, GIP (gastric inhibitory polypeptide), neurotensin, glicentin/glucagon-37 and PYY (peptide tyrosine tyrosine) are the main products of gastrointestinal endocrine cells; glucagon, CRF (corticotropin releasing factor), somatostatin, PP (pancreatic polypeptide) and GRF (growth hormone releasing factor), in addition to insulin, are produced in pancreatic islet cells; bombesin-related peptides are the main markers of pulmonary endocrine cells; calcitonin and CGRP (calcitonin gene-related peptide) occur in thyroid and extrathyroid C cells; ACTH and endorphins in anterior and intermediate lobe pituitary cells, alpha-MSH and CLIP (corticotropin-like intermediate lobe peptide) in intermediate lobe cells; met- and leu-enkephalins and related peptides in adrenal medullary and paraganglionic cells as well as in some gut (enterochromaffin) cells; NPY (neuropeptide Y) in adrenaline-type adrenal medullary cells, etc.. Both tissue-appropriate and tissue-inappropriate regulatory peptides are produced by endocrine tumours, with inappropriate peptides mostly produced by malignant tumours.

  6. Human periodontal ligament stem cells repair mental nerve injury*

    Institute of Scientific and Technical Information of China (English)

    Bohan Li; Hun-Jong Jung; Soung-Min Kim; Myung-Jin Kim; Jeong Won Jahng; Jong-Ho Lee

    2013-01-01

    Human periodontal ligament stem cells are easily accessible and can differentiate into Schwann cells. We hypothesized that human periodontal ligament stem cells can be used as an alternative source for the autologous Schwann cells in promoting the regeneration of injured peripheral nerve. To validate this hypothesis, human periodontal ligament stem cells (1 × 106) were injected into the crush-injured left mental nerve in rats. Simultaneously, autologous Schwann cells (1 × 106) and PBS were also injected as controls. Real-time reverse transcriptase polymerase chain reaction showed that at 5 days after injection, mRNA expression of low affinity nerve growth factor receptor was sig-nificantaly increased in the left trigeminal ganglion of rats with mental nerve injury. Sensory tests, histomorphometric evaluation and retrograde labeling demonstrated that at 2 and 4 weeks after in-jection, sensory function was significantly improved, the numbers of retrograde labeled sensory neurons and myelinated axons were significantly increased, and human periodontal ligament stem cells and autologous Schwann cells exhibited similar therapeutic effects. These findings suggest that transplantation of human periodontal ligament stem cells show a potential value in repair of mental nerve injury.

  7. Constriction of the buccal branch of the facial nerve produces unilateral craniofacial allodynia.

    Science.gov (United States)

    Lewis, Susannah S; Grace, Peter M; Hutchinson, Mark R; Maier, Steven F; Watkins, Linda R

    2016-12-18

    Despite pain being a sensory experience, studies of spinal cord ventral root damage have demonstrated that motor neuron injury can induce neuropathic pain. Whether injury of cranial motor nerves can also produce nociceptive hypersensitivity has not been addressed. Herein, we demonstrate that chronic constriction injury (CCI) of the buccal branch of the facial nerve results in long-lasting, unilateral allodynia in the rat. An anterograde and retrograde tracer (3000MW tetramethylrhodamine-conjugated dextran) was not transported to the trigeminal ganglion when applied to the injury site, but was transported to the facial nucleus, indicating that this nerve branch is not composed of trigeminal sensory neurons. Finally, intracisterna magna injection of interleukin-1 (IL-1) receptor antagonist reversed allodynia, implicating the pro-inflammatory cytokine IL-1 in the maintenance of neuropathic pain induced by facial nerve CCI. These data extend the prior evidence that selective injury to motor axons can enhance pain to supraspinal circuits by demonstrating that injury of a facial nerve with predominantly motor axons is sufficient for neuropathic pain, and that the resultant pain has a neuroimmune component.

  8. Rapamycin promotes Schwann cell migration and nerve growth factor secretion

    OpenAIRE

    Liu, Fang; Zhang, Haiwei; Zhang, Kaiming; Wang, Xinyu; Li, Shipu; Yin, Yixia

    2014-01-01

    Rapamycin, similar to FK506, can promote neural regeneration in vitro. We assumed that the mechanisms of action of rapamycin and FK506 in promoting peripheral nerve regeneration were similar. This study compared the effects of different concentrations of rapamycin and FK506 on Schwann cells and investigated effects and mechanisms of rapamycin on improving peripheral nerve regeneration. Results demonstrated that the lowest rapamycin concentration (1.53 nmol/L) more significantly promoted Schwa...

  9. Peripheral nerve extract effects on mesenchymal cells.

    Science.gov (United States)

    Dietz, F R; Mukhopadhyay, B; Becker, G; Daniels, K; Solursh, M

    1996-01-01

    Several common congenital limb disorders are characterized by normal tissue differentiation but abnormal somatic growth. These include: idiopathic clubfoot, idiopathic leg length discrepancy, hemi-atrophy and hemi-hypertrophy. Both clinical and research studies have suggested that peripheral nerves may be important in regulating somatic growth of limb tissues. To investigate the hypothesis that peripheral nerves convey trophic substances to mesenchymal tissues that are involved in the regulation of growth, we developed an in vitro assay to assess the effect of fractions of peripheral nerve on myoblast and chondroblast growth and differentiation in a mammalian (rat) system. Whole rat sciatic nerve extract was fractionated by ammonium sulfate precipitation and by affinity chromatography. Concavalin A chromatography resolved whole nerve extract into a glycoprotein and a non-glycoprotein fraction. Serial ammonium sulfate precipitation yielded three pellet fractions designated as 35%, 70%, and 100% pellets; corresponding to ammonium sulfate concentrations of 0 to 35%, 35 to 70%, and 70 to 100% saturation, respectively. Dialyzed solutions of these pellets as well as the fractions from Concavalin A chromatography were assayed for biological activity in micromass cultures of rat limb bud mesenchyme, which allowed assessment of both myoblast and chondroblast stimulation. Stimulation of protein synthesis and myoblast proliferation (as measured by MF20 staining) occurred with both 70% and 100% ammonium sulfate fractions. Stimulation of chondroblasts (as measured by the number of alcian blue staining nodules) occurred with the 35% and 100% fractions. The glycoprotein fraction from the affinity chromatography stimulated protein synthesis and myoblast proliferation and inhibited chondroblast development. Stimulation of chondroblasts was seen with the non-glycoprotein fraction. No effect on protein synthesis, myoblast proliferation or chondroblast proliferation was found in

  10. Electrical stimulation of nerve cells using conductive nanofibrous scaffolds for nerve tissue engineering.

    Science.gov (United States)

    Ghasemi-Mobarakeh, Laleh; Prabhakaran, Molamma P; Morshed, Mohammad; Nasr-Esfahani, Mohammad Hossein; Ramakrishna, Seeram

    2009-11-01

    Fabrication of scaffolds with suitable chemical, mechanical, and electrical properties is critical for the success of nerve tissue engineering. Electrical stimulation was directly applied to electrospun conductive nanofibrous scaffolds to enhance the nerve regeneration process. In the present study, electrospun conductive nanofibers were prepared by mixing 10 and 15 wt% doped polyaniline (PANI) with poly (epsilon-caprolactone)/gelatin (PG) (70:30) solution (PANI/PG) by electrospinning. The fiber diameter, pore size, hydrophilicity, tensile properties, conductivity, Fourier transform infrared (FTIR), and X-ray photoelectron spectroscopy spectra of nanofibers were determined, and the in vitro biodegradability of the different nanofibrous scaffolds was also evaluated. Nanofibrous scaffolds containing 15% PANI was found to exhibit the most balanced properties to meet all the required specifications for electrical stimulation for its enhanced conductivity and is used for in vitro culture and electrical stimulation of nerve stem cells. 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay and scanning electron microscopy results showed that conductive nanofibrous scaffolds are suitable substrates for the attachment and proliferation of nerve stem cells. Electrical stimulation through conductive nanofibrous PANI/PG scaffolds showed enhanced cell proliferation and neurite outgrowth compared to the PANI/PG scaffolds that were not subjected to electrical stimulation.

  11. Expression changes of nerve cell adhesion molecules L1 and semaphorin 3A after peripheral nerve injury

    OpenAIRE

    Qian-ru He; Meng Cong; Qing-zhong Chen; Ya-feng Sheng; Jian Li; Qi Zhang; Fei Ding; Yan-pei Gong

    2016-01-01

    The expression of nerve cell adhesion molecule L1 in the neuronal growth cone of the central nervous system is strongly associated with the direction of growth of the axon, but its role in the regeneration of the peripheral nerve is still unknown. This study explored the problem in a femoral nerve section model in rats. L1 and semaphorin 3A mRNA and protein expressions were measured over the 4-week recovery period. Quantitative polymerase chain reaction showed that nerve cell adhesion molecul...

  12. In vivo MRI monitoring nerve regeneration of acute peripheral nerve traction injury following mesenchymal stem cell transplantation

    Energy Technology Data Exchange (ETDEWEB)

    Duan, Xiao-Hui, E-mail: duanxiaohui-128@163.com [Department of Radiology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, No. 107 Yanjiang Road West, Guangzhou 510120, Guangdong (China); Cheng, Li-Na, E-mail: kobe10716@163.com [Department of Radiology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, No. 107 Yanjiang Road West, Guangzhou 510120, Guangdong (China); Zhang, Fang, E-mail: xinxin110007@yahoo.com.cn [Department of Radiology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, No. 107 Yanjiang Road West, Guangzhou 510120, Guangdong (China); Liu, Jun, E-mail: docliujun@hotmail.com [Department of Neurology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, No. 107 Yanjiang Road West, Guangzhou 510120, Guangdong (China); Guo, Ruo-Mi, E-mail: guoruomi-521@163.com [Department of Radiology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, No. 107 Yanjiang Road West, Guangzhou 510120, Guangdong (China); Zhong, Xiao-Mei, E-mail: enough300@yahoo.com.cn [Department of Radiology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, No. 107 Yanjiang Road West, Guangzhou 510120, Guangdong (China); Wen, Xue-Hua, E-mail: xuehuasuqian@126.com [Department of Radiology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, No. 107 Yanjiang Road West, Guangzhou 510120, Guangdong (China); Shen, Jun, E-mail: junshenjun@hotmail.com [Department of Radiology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, No. 107 Yanjiang Road West, Guangzhou 510120, Guangdong (China)

    2012-09-15

    Objective: To assess the continuous process of nerve regeneration in acute peripheral nerve traction injury treated with mesenchymal stem cells (MSCs) transplantation using MRI. Materials and methods: 1 week after acute nerve traction injury was established in the sciatic nerve of 48 New Zealand white rabbits, 5 × 10{sup 5} MSCs and vehicle alone were grafted to the acutely distracted sciatic nerves each in 24 animals. Serial MRI and T1 and T2 measurements of the injured nerves were performed with a 1.5-T scanner and functional recovery was recorded over a 10-week follow-up period, with histological assessments performed at regular intervals. Results: Compared with vehicle control, nerves grafted with MSCs had better functional recovery and showed improved nerve regeneration, with a sustained increase of T1 and T2 values during the phase of regeneration. Conclusion: MRI could be used to monitor the enhanced nerve regeneration in acute peripheral nerve traction injury treated with MSC transplantation, reflected by a prolonged increase in T1 and T2 values of the injured nerves.

  13. A Silk Fibroin/Collagen Nerve Scaffold Seeded with a Co-Culture of Schwann Cells and Adipose-Derived Stem Cells for Sciatic Nerve Regeneration.

    Directory of Open Access Journals (Sweden)

    Yunqiang Xu

    Full Text Available As a promising alternative to autologous nerve grafts, tissue-engineered nerve grafts have been extensively studied as a way to bridge peripheral nerve defects and guide nerve regeneration. The main difference between autogenous nerve grafts and tissue-engineered nerve grafts is the regenerative microenvironment formed by the grafts. If an appropriate regenerative microenvironment is provided, the repair of a peripheral nerve is feasible. In this study, to mimic the body's natural regenerative microenvironment closely, we co-cultured Schwann cells (SCs and adipose-derived stem cells (ADSCs as seed cells and introduced them into a silk fibroin (SF/collagen scaffold to construct a tissue-engineered nerve conduit (TENC. Twelve weeks after the three different grafts (plain SF/collagen scaffold, TENC, and autograft were transplanted to bridge 1-cm long sciatic nerve defects in rats, a series of electrophysiological examinations and morphological analyses were performed to evaluate the effect of the tissue-engineered nerve grafts on peripheral nerve regeneration. The regenerative outcomes showed that the effect of treatment with TENCs was similar to that with autologous nerve grafts but superior to that with plain SF/collagen scaffolds. Meanwhile, no experimental animals had inflammation around the grafts. Based on this evidence, our findings suggest that the TENC we developed could improve the regenerative microenvironment and accelerate nerve regeneration compared to plain SF/collagen and may serve as a promising strategy for peripheral nerve repair.

  14. A Silk Fibroin/Collagen Nerve Scaffold Seeded with a Co-Culture of Schwann Cells and Adipose-Derived Stem Cells for Sciatic Nerve Regeneration.

    Science.gov (United States)

    Xu, Yunqiang; Zhang, Zhenhui; Chen, Xuyi; Li, Ruixin; Li, Dong; Feng, Shiqing

    2016-01-01

    As a promising alternative to autologous nerve grafts, tissue-engineered nerve grafts have been extensively studied as a way to bridge peripheral nerve defects and guide nerve regeneration. The main difference between autogenous nerve grafts and tissue-engineered nerve grafts is the regenerative microenvironment formed by the grafts. If an appropriate regenerative microenvironment is provided, the repair of a peripheral nerve is feasible. In this study, to mimic the body's natural regenerative microenvironment closely, we co-cultured Schwann cells (SCs) and adipose-derived stem cells (ADSCs) as seed cells and introduced them into a silk fibroin (SF)/collagen scaffold to construct a tissue-engineered nerve conduit (TENC). Twelve weeks after the three different grafts (plain SF/collagen scaffold, TENC, and autograft) were transplanted to bridge 1-cm long sciatic nerve defects in rats, a series of electrophysiological examinations and morphological analyses were performed to evaluate the effect of the tissue-engineered nerve grafts on peripheral nerve regeneration. The regenerative outcomes showed that the effect of treatment with TENCs was similar to that with autologous nerve grafts but superior to that with plain SF/collagen scaffolds. Meanwhile, no experimental animals had inflammation around the grafts. Based on this evidence, our findings suggest that the TENC we developed could improve the regenerative microenvironment and accelerate nerve regeneration compared to plain SF/collagen and may serve as a promising strategy for peripheral nerve repair.

  15. Morphological changes in nerve cells during normal aging.

    Science.gov (United States)

    Pannese, Ennio

    2011-06-01

    During normal aging, widespread loss of nerve cells does not occur. Neuronal loss is limited to restricted regions of the nervous system and is slight (probably no more than 10%). The commonest age-related structural changes undergone by nerve cells are as follows: dendrites decrease in number and length and many dendritic spines are lost; axons decrease in number and their myelin sheaths may become less compact and undergo segmental demyelination followed by remyelination; and significant loss of synapses occurs. These changes probably make a significant contribution to the behavioral impairment and cognitive decline that often accompany normal aging.

  16. Nerve agent analogues that produce authentic soman, sarin, tabun, and cyclohexyl methylphosphonate-modified human butyrylcholinesterase.

    Science.gov (United States)

    Gilley, Cynthia; MacDonald, Mary; Nachon, Florian; Schopfer, Lawrence M; Zhang, Jun; Cashman, John R; Lockridge, Oksana

    2009-10-01

    The goal was to test 14 nerve agent model compounds of soman, sarin, tabun, and cyclohexyl methylphosphonofluoridate (GF) for their suitability as substitutes for true nerve agents. We wanted to know whether the model compounds would form the identical covalent adduct with human butyrylcholinesterase that is produced by reaction with true nerve agents. Nerve agent model compounds containing thiocholine or thiomethyl in place of fluorine or cyanide were synthesized as Sp and Rp stereoisomers. Purified human butyrylcholinesterase was treated with a 45-fold molar excess of nerve agent analogue at pH 7.4 for 17 h at 21 degrees C. The protein was denatured by boiling and was digested with trypsin. Aged and nonaged active site peptide adducts were quantified by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry of the tryptic digest mixture. The active site peptides were isolated by HPLC and analyzed by MALDI-TOF-TOF mass spectrometry. Serine 198 of butyrylcholinesterase was covalently modified by all 14 compounds. Thiocholine was the leaving group in all compounds that had thiocholine in place of fluorine or cyanide. Thiomethyl was the leaving group in the GF thiomethyl compounds. However, sarin thiomethyl compounds released either thiomethyl or isopropyl, while soman thiomethyl compounds released either thiomethyl or pinacolyl. Thiocholine compounds reacted more rapidly with butyrylcholinesterase than thiomethyl compounds. Labeling with the model compounds resulted in aged adducts that had lost the O-alkyl group (O-ethyl for tabun, O-cyclohexyl for GF, isopropyl for sarin, and pinacolyl for soman) in addition to the thiocholine or thiomethyl group. The nerve agent model compounds containing thiocholine and the GF thiomethyl analogue were found to be suitable substitutes for true soman, sarin, tabun, and GF in terms of the adduct that they produced with human butyrylcholinesterase. However, the soman and sarin thiomethyl compounds

  17. Coordinated, multi-joint, fatigue-resistant feline stance produced with intrafascicular hind limb nerve stimulation

    Science.gov (United States)

    Normann, R. A.; Dowden, B. R.; Frankel, M. A.; Wilder, A. M.; Hiatt, S. D.; Ledbetter, N. M.; Warren, D. A.; Clark, G. A.

    2012-04-01

    The production of graceful skeletal movements requires coordinated activation of multiple muscles that produce torques around multiple joints. The work described herein is focused on one such movement, stance, that requires coordinated activation of extensor muscles acting around the hip, knee and ankle joints. The forces evoked in these muscles by external stimulation all have a complex dependence on muscle length and shortening velocities, and some of these muscles are biarticular. In order to recreate sit-to-stand maneuvers in the anesthetized feline, we excited the hind limb musculature using intrafascicular multielectrode stimulation (IFMS) of the muscular branch of the sciatic nerve, the femoral nerve and the main branch of the sciatic nerve. Stimulation was achieved with either acutely or chronically implanted Utah Slanted Electrode Arrays (USEAs) via subsets of electrodes (1) that activated motor units in the extensor muscles of the hip, knee and ankle joints, (2) that were able to evoke large extension forces and (3) that manifested minimal coactivation of the targeted motor units. Three hind limb force-generation strategies were investigated, including sequential activation of independent motor units to increase force, and interleaved or simultaneous IFMS of three sets of six or more USEA electrodes that excited the hip, knee and ankle extensors. All force-generation strategies evoked stance, but the interleaved IFMS strategy also reduced muscle fatigue produced by repeated sit-to-stand maneuvers compared with fatigue produced by simultaneous activation of different motor neuron pools. These results demonstrate the use of interleaved IFMS as a means to recreate coordinated, fatigue-resistant multi-joint muscle forces in the unilateral hind limb. This muscle activation paradigm could provide a promising neuroprosthetic approach for the restoration of sit-to-stand transitions in individuals who are paralyzed by spinal cord injury, stroke or disease.

  18. In vivo integration of poly(ε-caprolactone)/gelatin nanofibrous nerve guide seeded with teeth derived stem cells for peripheral nerve regeneration.

    Science.gov (United States)

    Beigi, Mohammad-Hossein; Ghasemi-Mobarakeh, Laleh; Prabhakaran, Molamma P; Karbalaie, Khadijeh; Azadeh, Hamid; Ramakrishna, Seeram; Baharvand, Hossein; Nasr-Esfahani, Mohammad-Hossein

    2014-12-01

    Artificial nanofiber nerve guides have gained huge interest in bridging nerve gaps and associated peripheral nerve regeneration due to its high surface area, flexibility and porous structure. In this study, electrospun poly (ε-caprolactone)/gelatin (PCL/Gel) nanofibrous mats were fabricated, rolled around a copper wire and fixed by medical grade adhesive to obtain a tubular shaped bio-graft, to bridge 10 mm sciatic nerve gap in in vivo rat models. Stem cells from human exfoliated deciduous tooth (SHED) were transplanted to the site of nerve injury through the nanofibrous nerve guides. In vivo experiments were performed in animal models after creating a sciatic nerve gap, such that the nerve gap was grafted using (i) nanofiber nerve guide (ii) nanofiber nerve guide seeded with SHED (iii) suturing, while an untreated nerve gap remained as the negative control. In vitro cell culture study was carried out for primary investigation of SHED-nanofiber interaction and its viability within the nerve guides after 2 and 16 weeks of implantation time. Walking track analysis, plantar test, electrophysiology and immunohistochemistry were performed to evaluate functional recovery during nerve regeneration. Vascularization was also investigated by hematoxilin/eosine (H&E) staining. Overall results showed that the SHED seeded on nanofibrous nerve guide could survive and promote axonal regeneration in rat sciatic nerves, whereby the biocompatible PCL/Gel nerve guide with cells can support axonal regeneration and could be a promising tissue engineered graft for peripheral nerve regeneration.

  19. Bridging long gap peripheral nerve injury using skeletal muscle-derived multipotent stem cells

    Institute of Scientific and Technical Information of China (English)

    Tetsuro Tamaki

    2014-01-01

    Long gap peripheral nerve injuries usually reulting in life-changing problems for patients. Skeletal muscle derived-multipotent stem cells (Sk-MSCs) can differentiate into Schwann and perineurial/endoneurial cells, vascular relating pericytes, and endothelial and smooth muscle cells in the damaged peripheral nerve niche. Application of the Sk-MSCs in the bridging conduit for repairing long nerve gap injury resulted favorable axonal regeneration, which showing supe-rior effects than gold standard therapy--healthy nerve autograft. This means that it does not need to sacriifce of healthy nerves or loss of related functions for repairing peripheral nerve injury.

  20. Human amniotic epithelial cell transplantation for the repair of injured brachial plexus nerve: evaluation of nerve viscoelastic properties

    Directory of Open Access Journals (Sweden)

    Hua Jin

    2015-01-01

    Full Text Available The transplantation of embryonic stem cells can effectively improve the creeping strength of nerves near an injury site in animals. Amniotic epithelial cells have similar biological properties as embryonic stem cells; therefore, we hypothesized that transplantation of amniotic epithelial cells can repair peripheral nerve injury and recover the creeping strength of the brachial plexus nerve. In the present study, a brachial plexus injury model was established in rabbits using the C 6 root avulsion method. A suspension of human amniotic epithelial cells was repeatedly injected over an area 4.0 mm lateral to the cephal and caudal ends of the C 6 brachial plexus injury site (1 × 10 6 cells/mL, 3 μL/injection, 25 injections immediately after the injury. The results showed that the decrease in stress and increase in strain at 7,200 seconds in the injured rabbit C 6 brachial plexus nerve were mitigated by the cell transplantation, restoring the viscoelastic stress relaxation and creep properties of the brachial plexus nerve. The forepaw functions were also significantly improved at 26 weeks after injury. These data indicate that transplantation of human amniotic epithelial cells can effectively restore the mechanical properties of the brachial plexus nerve after injury in rabbits and that viscoelasticity may be an important index for the evaluation of brachial plexus injury in animals.

  1. Human amniotic epithelial cell transplantation for the repair of injured brachial plexus nerve:evaluation of nerve viscoelastic properties

    Institute of Scientific and Technical Information of China (English)

    Hua Jin; Qi Yang; Feng Ji; Ya-jie Zhang; Yan Zhao; Min Luo

    2015-01-01

    The transplantation of embryonic stem cells can effectively improve the creeping strength of nerves near an injury site in animals. Amniotic epithelial cells have similar biological properties as em-bryonic stem cells; therefore, we hypothesized that transplantation of amniotic epithelial cells can repair peripheral nerve injury and recover the creeping strength of the brachial plexus nerve. In the present study, a brachial plexus injury model was established in rabbits using the C6root avulsion method. A suspension of human amniotic epithelial cells was repeatedly injected over an area 4.0 mm lateral to the cephal and caudal ends of the C6 brachial plexus injury site (1 × 106 cells/mL, 3μL/injection, 25 injections) immediately after the injury. The results showed that the decrease in stress and increase in strain at 7,200 seconds in the injured rabbit C6 brachial plexus nerve were mitigated by the cell transplantation, restoring the viscoelastic stress relaxation and creep properties of the brachial plexus nerve. The forepaw functions were also signiifcantly improved at 26 weeks after injury. These data indicate that transplantation of human amniotic epithelial cells can effec-tively restore the mechanical properties of the brachial plexus nerve after injury in rabbits and that viscoelasticity may be an important index for the evaluation of brachial plexus injury in animals.

  2. Tonsil-Derived Mesenchymal Stem Cells Differentiate into a Schwann Cell Phenotype and Promote Peripheral Nerve Regeneration.

    Science.gov (United States)

    Jung, Namhee; Park, Saeyoung; Choi, Yoonyoung; Park, Joo-Won; Hong, Young Bin; Park, Hyun Ho Choi; Yu, Yeonsil; Kwak, Geon; Kim, Han Su; Ryu, Kyung-Ha; Kim, Jae Kwang; Jo, Inho; Choi, Byung-Ok; Jung, Sung-Chul

    2016-11-09

    Schwann cells (SCs), which produce neurotropic factors and adhesive molecules, have been reported previously to contribute to structural support and guidance during axonal regeneration; therefore, they are potentially a crucial target in the restoration of injured nervous tissues. Autologous SC transplantation has been performed and has shown promising clinical results for treating nerve injuries and donor site morbidity, and insufficient production of the cells have been considered as a major issue. Here, we performed differentiation of tonsil-derived mesenchymal stem cells (T-MSCs) into SC-like cells (T-MSC-SCs), to evaluate T-MSC-SCs as an alternative to SCs. Using SC markers such as CAD19, GFAP, MBP, NGFR, S100B, and KROX20 during quantitative real-time PCR we detected the upregulation of NGFR, S100B, and KROX20 and the downregulation of CAD19 and MBP at the fully differentiated stage. Furthermore, we found myelination of axons when differentiated SCs were cocultured with mouse dorsal root ganglion neurons. The application of T-MSC-SCs to a mouse model of sciatic nerve injury produced marked improvements in gait and promoted regeneration of damaged nerves. Thus, the transplantation of human T-MSCs might be suitable for assisting in peripheral nerve regeneration.

  3. Skeletal muscle cell apoptosis following motornerve injury versus sensory nerve injury

    Institute of Scientific and Technical Information of China (English)

    Lei Zhao; Ruisheng Xu; Shenyang Jiang; Guangming Lü; Zhiqiang Yan; Junming Sun; Ling Wang; Ye Xue; Donglin Jiang

    2011-01-01

    Skeletal muscle atrophy inevitably occurs in denervated skeletal muscle, and cell apoptosis plays an important role in skeletal muscle atrophy and degeneration. The present study established rat models of simple nerve injury by transecting the ventral or dorsal spinal nerve root and observed rat skeletal muscle cell apoptosis following simple motor nerve injury versus simple sensory nerve injury. Following skeletal muscle denervation for 10 weeks, cell apoptosis was detected in skeletal muscle, which was accompanied by obvious changes in rat behavior and electrophysiological responses. In addition, changes in cross-sectional area and average gray-scale of motor endplates of the gastrocnemius muscle were analyzed following sciatic nerve injury and motor nerve injury.Cell nuclei in denervated skeletal muscle tissue were more densely arranged than in normal skeletal muscle tissue. Cell nuclei were most dense in the sciatic nerve injury group, followed by the motor nerve injury group and the sensory nerve injury group. Fas/Fast expression and the number of apoptotic cells increased in denervated skeletal muscle, and apoptosis-related changes were observed. These findings suggested that motor and sensory nerves provided trophic actions following skeletal muscle and motor nerve injury, resulting in a greater influence on skeletal muscle atrophy than sensory nerve injury. Therefore, reconstruction of motor nerves should be preferentially considered for treating denervation-induced skeletal muscle atrophy.

  4. Bridging sciatic nerve gap using tissue-engineered nerves constructed with neural tissue-committed stem cells derived from bone marrow

    Institute of Scientific and Technical Information of China (English)

    Zhiying Zhang; Congli Ren; Chuansen Zhang; Fang Liu; Liang Li

    2009-01-01

    BACKGROUND: Schwann cells are the most commonly used cells for tissue-engineered nerves. However, autologous Schwann cells are of limited use in a clinical context, and allogeneic Schwann cells induce immunological rejections. Cells that do not induce immunological rejections and that are relatively easy to acquire are urgently needed for transplantation.OBJECTIVE: To bridge sciatic nerve defects using tissue engineered nerves constructed with neural tissue-committed stem cells (NTCSCs) derived from bone marrow; to observe morphology and function of rat nerves following bridging; to determine the effect of autologous nerve transplantation, which serves as the gold standard for evaluating efficacy of tissue-engineered nerves.DESIGN, TIME AND SETTING: This randomized, controlled, animal experiment was performed in the Anatomical laboratory and Biomedical Institute of the Second Military Medical University of Chinese PLA between September 2004 and April 2006.MATERIALS: Five Sprague Dawley rats, aged 1 month and weighing 100-150 g, were used for cell culture. Sixty Sprague Dawiey rats aged 3 months and weighing 220-250 g, were used to establish neurological defect models. Nestin, neuron-specific enolase (NSE), glial fibrillary acidic protein (GFAP), and S-100 antibodies were provided by Santa Cruz Biotechnology, Inc., USA. Acellular nerve grafts were derived from dogs.METHODS: All rats, each with 1-cm gap created in the right sciatic nerve, were randomly assigned to three groups. Each group comprised 20 rats. Autograft nerve transplantation group: the severed 1-cm length nerve segment was reverted, but with the two ends exchanged; the proximal segment was sutured to the distal sciatic nerve stump and the distal segment to the proximal stump. Blank nerve scaffold transplantation group: a 1-cm length acellular nerve graft was used to bridge the sciatic nerve gap. NTCSC engineered nerve transplantation group: a 1-cm length acellular nerve graft, in which NTCSCs were

  5. Mesenchymal stem cells in a polycaprolactone conduit promote sciatic nerve regeneration and sensory neuron survival after nerve injury.

    Science.gov (United States)

    Frattini, Flávia; Lopes, Fatima Rosalina Pereira; Almeida, Fernanda Martins; Rodrigues, Rafaela Fintelman; Boldrini, Leonardo Cunha; Tomaz, Marcelo A; Baptista, Abrahão Fontes; Melo, Paulo A; Martinez, Ana Maria Blanco

    2012-10-01

    Despite the fact that the peripheral nervous system is able to regenerate after traumatic injury, the functional outcomes following damage are limited and poor. Bone marrow mesenchymal stem cells (MSCs) are multipotent cells that have been used in studies of peripheral nerve regeneration and have yielded promising results. The aim of this study was to evaluate sciatic nerve regeneration and neuronal survival in mice after nerve transection followed by MSC treatment into a polycaprolactone (PCL) nerve guide. The left sciatic nerve of C57BL/6 mice was transected and the nerve stumps were placed into a biodegradable PCL tube leaving a 3-mm gap between them; the tube was filled with MSCs obtained from GFP+ animals (MSC-treated group) or with a culture medium (Dulbecco's modified Eagle's medium group). Motor function was analyzed according to the sciatic functional index (SFI). After 6 weeks, animals were euthanized, and the regenerated sciatic nerve, the dorsal root ganglion (DRG), the spinal cord, and the gastrocnemius muscle were collected and processed for light and electron microscopy. A quantitative analysis of regenerated nerves showed a significant increase in the number of myelinated fibers in the group that received, within the nerve guide, stem cells. The number of neurons in the DRG was significantly higher in the MSC-treated group, while there was no difference in the number of motor neurons in the spinal cord. We also found higher values of trophic factors expression in MSC-treated groups, especially a nerve growth factor. The SFI revealed a significant improvement in the MSC-treated group. The gastrocnemius muscle showed an increase in weight and in the levels of creatine phosphokinase enzyme, suggesting an improvement of reinnervation and activity in animals that received MSCs. Immunohistochemistry documented that some GFP+ -transplanted cells assumed a Schwann-cell-like phenotype, as evidenced by their expression of the S-100 protein, a Schwann cell

  6. Human amniotic epithelial cells express specific markers of nerve cells and migrate along the nerve fibers in the corpus callosum

    Institute of Scientific and Technical Information of China (English)

    Zhiyuan Wu; Guozhen Hui; Yi Lu; Tianjin Liu; Qin Huang; Lihe Guo

    2012-01-01

    Human amniotic epithelial cells were isolated from a piece of fresh amnion. Using immunocytochemical methods, we investigated the expression of neuronal phenotypes (microtubule-associated protein-2, glial fibrillary acidic protein and nestin) in human amniotic epithelial cells. The conditioned medium of human amniotic epithelial cells promoted the growth and proliferation of rat glial cells cultured in vitro, and this effect was dose-dependent. Human amniotic epithelial cells were further transplanted into the corpus striatum of healthy adult rats and the grafted cells could integrate with the host and migrate 1-2 mm along the nerve fibers in corpus callosum. Our experimental findings indicate that human amniotic epithelial cells may be a new kind of seed cells for use in neurograft.

  7. Dendritic cell-nerve clusters are sites of T cell proliferation in allergic airway inflammation.

    Science.gov (United States)

    Veres, Tibor Z; Shevchenko, Marina; Krasteva, Gabriela; Spies, Emma; Prenzler, Frauke; Rochlitzer, Sabine; Tschernig, Thomas; Krug, Norbert; Kummer, Wolfgang; Braun, Armin

    2009-03-01

    Interactions between T cells and dendritic cells in the airway mucosa precede secondary immune responses to inhaled antigen. The purpose of this study was to identify the anatomical locations where dendritic cell-T cell interactions occur, resulting in T cells activation by dendritic cells. In a mouse model of allergic airway inflammation, we applied whole-mount immunohistology and confocal microscopy to visualize dendritic cells and T cells together with nerves, epithelium, and smooth muscle in three dimensions. Proliferating T cells were identified by the detection of the incorporation of the nucleotide analogue 5-ethynyl-2'-deoxyuridine into the DNA. We developed a novel quantification method that enabled the accurate determination of cell-cell contacts in a semi-automated fashion. Dendritic cell-T cell interactions occurred beneath the smooth muscle layer, but not in the epithelium. Approximately 10% of the dendritic cells were contacted by nerves, and up to 4% of T cells formed clusters with these dendritic cells. T cells that were clustered with nerve-contacting dendritic cells proliferated only in the airways of mice with allergic inflammation but not in the airways of negative controls. Taken together, these results suggest that during the secondary immune response, sensory nerves influence dendritic cell-driven T cell activation in the airway mucosa.

  8. Chronic experimental myocardial infarction produces antinatriuresis by a renal nerve-dependent mechanism

    Directory of Open Access Journals (Sweden)

    Souza D.R.B.

    2004-01-01

    Full Text Available The present study focused on the role of sympathetic renal nerve activity, in mediating congestive heart failure-induced sodium retention following experimental chronic myocardial infarction. Groups of male Wistar rats (240-260 g were studied: sham-operated coronary ligation (CON3W, N = 11, coronary ligation and sham-operated renal denervation (INF3W, N = 19, 3 weeks of coronary ligation and sympathetic renal nerve denervation (INF3WDX, N = 6, sham-operated coronary ligation (N = 7, and 16 weeks of coronary ligation (INF16W, N = 7. An acute experimental protocol was used in which the volume overload (VO; 5% of body weight was applied for 30 min after the equilibration period of continuous iv infusion of saline. Compared to control levels, VO produced an increase (P < 0.01, ANOVA in urine flow rate (UFR; 570% and urinary sodium excretion (USE; 1117% in CON3W. VO induced a smaller increase (P < 0.01 in USE (684% in INF3W. A similar response was also observed in INF16W. In INF3WDX, VO produced an immediate and large increase (P < 0.01 in UFR (547% and USE (1211%. Similarly, in INF3W VO increased (P < 0.01 UFR (394% and USE (894%. Compared with INF3W, VO induced a higher (P < 0.01 USE in INF3WDX, whose values were similar to those for CON3W. These results suggest that renal sympathetic activity may be involved in sodium retention induced by congestive heart failure. This premise is supported by the observation that in bilaterally renal denervated INF3WDX rats myocardial infarction was unable to reduce volume expansion-induced natriuresis. However, the mechanism involved in urinary volume regulation seems to be insensitive to the factors that alter natriuresis.

  9. Nerve growth factor promotes in vitro proliferation of neural stem cells from tree shrews.

    Science.gov (United States)

    Xiong, Liu-Lin; Chen, Zhi-Wei; Wang, Ting-Hua

    2016-04-01

    Neural stem cells promote neuronal regeneration and repair of brain tissue after injury, but have limited resources and proliferative ability in vivo. We hypothesized that nerve growth factor would promote in vitro proliferation of neural stem cells derived from the tree shrews, a primate-like mammal that has been proposed as an alternative to primates in biomedical translational research. We cultured neural stem cells from the hippocampus of tree shrews at embryonic day 38, and added nerve growth factor (100 μg/L) to the culture medium. Neural stem cells from the hippocampus of tree shrews cultured without nerve growth factor were used as controls. After 3 days, fluorescence microscopy after DAPI and nestin staining revealed that the number of neurospheres and DAPI/nestin-positive cells was markedly greater in the nerve growth factor-treated cells than in control cells. These findings demonstrate that nerve growth factor promotes the proliferation of neural stem cells derived from tree shrews.

  10. Nerve growth factor promotes in vitro proliferation of neural stem cells from tree shrews

    Institute of Scientific and Technical Information of China (English)

    Liu-lin Xiong; Zhi-wei Chen; Ting-hua Wang

    2016-01-01

    Neural stem cells promote neuronal regeneration and repair of brain tissue after injury, but have limited resources and proliferative ability in vivo. We hypothesized that nerve growth factor would promotein vitro proliferation of neural stem cells derived from the tree shrews, a primate-like mammal that has been proposed as an alternative to primates in biomedical translational research. We cultured neural stem cells from the hippocampus of tree shrews at embryonic day 38, and added nerve growth factor (100 μg/L) to the culture medium. Neural stem cells from the hippocampus of tree shrews cultured without nerve growth factor were used as controls. After 3 days, lfuorescence mi-croscopy after DAPI and nestin staining revealed that the number of neurospheres and DAPI/nestin-positive cells was markedly greater in the nerve growth factor-treated cells than in control cells. These ifndings demonstrate that nerve growth factor promotes the proliferation of neural stem cells derived from tree shrews.

  11. Characterization and Schwann Cell Seeding of up to 15.0 cm Long Spider Silk Nerve Conduits for Reconstruction of Peripheral Nerve Defects

    Directory of Open Access Journals (Sweden)

    Tim Kornfeld

    2016-11-01

    Full Text Available Nerve reconstruction of extended nerve defect injuries still remains challenging with respect to therapeutic options. The gold standard in nerve surgery is the autologous nerve graft. Due to the limitation of adequate donor nerves, surgical alternatives are needed. Nerve grafts made out of either natural or artificial materials represent this alternative. Several biomaterials are being explored and preclinical and clinical applications are ongoing. Unfortunately, nerve conduits with successful enhancement of axonal regeneration for nerve defects measuring over 4.0 cm are sparse and no conduits are available for nerve defects extending to 10.0 cm. In this study, spider silk nerve conduits seeded with Schwann cells were investigated for in vitro regeneration on defects measuring 4.0 cm, 10.0 cm and 15.0 cm in length. Schwann cells (SCs were isolated, cultured and purified. Cell purity was determined by immunofluorescence. Nerve grafts were constructed out of spider silk from Nephila edulis and decellularized ovine vessels. Finally, spider silk implants were seeded with purified Schwann cells. Cell attachment was observed within the first hour. After 7 and 21 days of culture, immunofluorescence for viability and determination of Schwann cell proliferation and migration throughout the conduits was performed. Analyses revealed that SCs maintained viable (>95% throughout the conduits independent of construct length. SC proliferation on the spider silk was determined from day 7 to day 21 with a proliferation index of 49.42% arithmetically averaged over all conduits. This indicates that spider silk nerve conduits represent a favorable environment for SC attachment, proliferation and distribution over a distance of least 15.0 cm in vitro. Thus spider silk nerve implants are a highly adequate biomaterial for nerve reconstruction.

  12. Characterization and Schwann Cell Seeding of up to 15.0 cm Long Spider Silk Nerve Conduits for Reconstruction of Peripheral Nerve Defects.

    Science.gov (United States)

    Kornfeld, Tim; Vogt, Peter M; Bucan, Vesna; Peck, Claas-Tido; Reimers, Kerstin; Radtke, Christine

    2016-11-30

    Nerve reconstruction of extended nerve defect injuries still remains challenging with respect to therapeutic options. The gold standard in nerve surgery is the autologous nerve graft. Due to the limitation of adequate donor nerves, surgical alternatives are needed. Nerve grafts made out of either natural or artificial materials represent this alternative. Several biomaterials are being explored and preclinical and clinical applications are ongoing. Unfortunately, nerve conduits with successful enhancement of axonal regeneration for nerve defects measuring over 4.0 cm are sparse and no conduits are available for nerve defects extending to 10.0 cm. In this study, spider silk nerve conduits seeded with Schwann cells were investigated for in vitro regeneration on defects measuring 4.0 cm, 10.0 cm and 15.0 cm in length. Schwann cells (SCs) were isolated, cultured and purified. Cell purity was determined by immunofluorescence. Nerve grafts were constructed out of spider silk from Nephila edulis and decellularized ovine vessels. Finally, spider silk implants were seeded with purified Schwann cells. Cell attachment was observed within the first hour. After 7 and 21 days of culture, immunofluorescence for viability and determination of Schwann cell proliferation and migration throughout the conduits was performed. Analyses revealed that SCs maintained viable (>95%) throughout the conduits independent of construct length. SC proliferation on the spider silk was determined from day 7 to day 21 with a proliferation index of 49.42% arithmetically averaged over all conduits. This indicates that spider silk nerve conduits represent a favorable environment for SC attachment, proliferation and distribution over a distance of least 15.0 cm in vitro. Thus spider silk nerve implants are a highly adequate biomaterial for nerve reconstruction.

  13. Construction of nerve guide conduits from cellulose/soy protein composite membranes combined with Schwann cells and pyrroloquinoline quinone for the repair of peripheral nerve defect

    Energy Technology Data Exchange (ETDEWEB)

    Luo, Lihua [Department of Biomedical Engineering, School of Basic Medical Sciences, Wuhan University, Wuhan 430071 (China); Center of Molecular Medicine, School of Medicine, Hubei University of Arts and Sciences, Xiangyang 441053 (China); Gan, Li; Liu, Yongming; Tian, Weiqun; Tong, Zan [Department of Biomedical Engineering, School of Basic Medical Sciences, Wuhan University, Wuhan 430071 (China); Wang, Xiong; Huselstein, Celine [Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), UMR 7365 CNRS – Université de Lorraine, Biopôle, 54500 Vandoeuvre-lès-Nancy (France); Chen, Yun, E-mail: yunchen@whu.edu.cn [Department of Biomedical Engineering, School of Basic Medical Sciences, Wuhan University, Wuhan 430071 (China)

    2015-02-20

    Regeneration and functional reconstruction of peripheral nerve defects remained a significant clinical challenge. Nerve guide conduits, with seed cells or neurotrophic factors (NTFs), had been widely used to improve the repair and regeneration of injured peripheral nerve. Pyrroloquinoline quinone (PQQ) was an antioxidant that can stimulate nerve growth factors (NGFs) synthesis and accelerate the Schwann cells (SCs) proliferation and growth. In present study, three kinds of nerve guide conduits were constructed: one from cellulose/SPI hollow tube (CSC), another from CSC combined with SCs (CSSC), and the third one from CSSC combined with PQQ (CSSPC), respectively. And then they were applied to bridge and repair the sciatic nerve defect in rats, using autograft as control. Effects of different nerve guide conduits on the nerve regeneration were comparatively evaluated by general analysis, sciatic function index (SFI) and histological analysis (HE and TEM). Newly-formed regenerative nerve fibers were observed and running through the transparent nerve guide conduits 12 weeks after surgery. SFI results indicated that the reconstruction of motor function in CSSPC group was better than that in CSSC and CSC groups. HE images from the cross-sections and longitudinal-sections of the harvested regenerative nerve indicated that regenerative nerve fibers had been formed and accompanied with new blood vessels and matrix materials in the conduits. TEM images also showed that lots of fresh myelinated and non-myelinated nerve fibers had been formed. Parts of vacuolar, swollen and abnormal axons occurred in CSC and CSSC groups, while the vacuolization and swell of axons was the least serious in CSSPC group. These results indicated that CSSPC group had the most ability to repair and reconstruct the nerve structure and functions due to the comprehensive contributions from hollow CSC tube, SCs and PQQ. As a result, the CSSPC may have the potential for the applications as nerve guide

  14. Construction of nerve guide conduits from cellulose/soy protein composite membranes combined with Schwann cells and pyrroloquinoline quinone for the repair of peripheral nerve defect.

    Science.gov (United States)

    Luo, Lihua; Gan, Li; Liu, Yongming; Tian, Weiqun; Tong, Zan; Wang, Xiong; Huselstein, Celine; Chen, Yun

    2015-02-20

    Regeneration and functional reconstruction of peripheral nerve defects remained a significant clinical challenge. Nerve guide conduits, with seed cells or neurotrophic factors (NTFs), had been widely used to improve the repair and regeneration of injured peripheral nerve. Pyrroloquinoline quinone (PQQ) was an antioxidant that can stimulate nerve growth factors (NGFs) synthesis and accelerate the Schwann cells (SCs) proliferation and growth. In present study, three kinds of nerve guide conduits were constructed: one from cellulose/SPI hollow tube (CSC), another from CSC combined with SCs (CSSC), and the third one from CSSC combined with PQQ (CSSPC), respectively. And then they were applied to bridge and repair the sciatic nerve defect in rats, using autograft as control. Effects of different nerve guide conduits on the nerve regeneration were comparatively evaluated by general analysis, sciatic function index (SFI) and histological analysis (HE and TEM). Newly-formed regenerative nerve fibers were observed and running through the transparent nerve guide conduits 12 weeks after surgery. SFI results indicated that the reconstruction of motor function in CSSPC group was better than that in CSSC and CSC groups. HE images from the cross-sections and longitudinal-sections of the harvested regenerative nerve indicated that regenerative nerve fibers had been formed and accompanied with new blood vessels and matrix materials in the conduits. TEM images also showed that lots of fresh myelinated and non-myelinated nerve fibers had been formed. Parts of vacuolar, swollen and abnormal axons occurred in CSC and CSSC groups, while the vacuolization and swell of axons was the least serious in CSSPC group. These results indicated that CSSPC group had the most ability to repair and reconstruct the nerve structure and functions due to the comprehensive contributions from hollow CSC tube, SCs and PQQ. As a result, the CSSPC may have the potential for the applications as nerve guide

  15. Sciatic nerve repair with tissue engineered nerve: Olfactory ensheathing cells seeded poly(lactic-co-glygolic acid conduit in an animal model

    Directory of Open Access Journals (Sweden)

    C W Tan

    2013-01-01

    Full Text Available Background and Aim: Synthetic nerve conduits have been sought for repair of nerve defects as the autologous nerve grafts causes donor site morbidity and possess other drawbacks. Many strategies have been investigated to improve nerve regeneration through synthetic nerve guided conduits. Olfactory ensheathing cells (OECs that share both Schwann cell and astrocytic characteristics have been shown to promote axonal regeneration after transplantation. The present study was driven by the hypothesis that tissue-engineered poly(lactic-co-glycolic acid (PLGA seeded with OECs would improve peripheral nerve regeneration in a long sciatic nerve defect. Materials and Methods: Sciatic nerve gap of 15 mm was created in six adult female Sprague-Dawley rats and implanted with PLGA seeded with OECs. The nerve regeneration was assessed electrophysiologically at 2, 4 and 6 weeks following implantation. Histopathological examination, scanning electron microscopic (SEM examination and immunohistochemical analysis were performed at the end of the study. Results: Nerve conduction studies revealed a significant improvement of nerve conduction velocities whereby the mean nerve conduction velocity increases from 4.2 ΁ 0.4 m/s at week 2 to 27.3 ΁ 5.7 m/s at week 6 post-implantation ( P < 0.0001. Histological analysis revealed presence of spindle-shaped cells. Immunohistochemical analysis further demonstrated the expression of S100 protein in both cell nucleus and the cytoplasm in these cells, hence confirming their Schwann-cell-like property. Under SEM, these cells were found to be actively secreting extracellular matrix. Conclusion: Tissue-engineered PLGA conduit seeded with OECs provided a permissive environment to facilitate nerve regeneration in a small animal model.

  16. Innexin gap junctions in nerve cells coordinate spontaneous contractile behavior in Hydra polyps

    KAUST Repository

    Takaku, Yasuharu

    2014-01-07

    Nerve cells and spontaneous coordinated behavior first appeared near the base of animal evolution in the common ancestor of cnidarians and bilaterians. Experiments on the cnidarian Hydra have demonstrated that nerve cells are essential for this behavior, although nerve cells in Hydra are organized in a diffuse network and do not form ganglia. Here we show that the gap junction protein innexin-2 is expressed in a small group of nerve cells in the lower body column of Hydra and that an anti-innexin-2 antibody binds to gap junctions in the same region. Treatment of live animals with innexin-2 antibody eliminates gap junction staining and reduces spontaneous body column contractions. We conclude that a small subset of nerve cells, connected by gap junctions and capable of synchronous firing, act as a pacemaker to coordinate the contraction of the body column in the absence of ganglia.

  17. Ligation of mouse L4 and L5 spinal nerves produces robust allodynia without major motor function deficit.

    Science.gov (United States)

    Ye, Gui-Lan; Savelieva, Katerina V; Vogel, Peter; Baker, Kevin B; Mason, Sara; Lanthorn, Thomas H; Rajan, Indrani

    2015-01-01

    Spinal nerve L5/L6 ligation (SNL) in rats has become the standard for mechanistic studies of peripheral neuropathy and screening for novel analgesics. Conventional SNL in our hybrid mice resulted in a wide range of allodynia. Anatomical evaluation indicated that a variable number of lumbar vertebrae existed, resulting in L4/L5 or L5/L6 being ligated. Surprisingly, L4/L5 ligation did not result in ipsilateral hind limb paralysis and produced robust allodynia. Following a recent report that the mouse L4 neural segment is homologous with rat L5 we generated L4, L5 or both L4 and L5 (L4/L5) ligations in C57 mice after establishing a modified set of surgical landmarks. In contrast to rats, L4 ligation in these mice did not result in hind limb paralysis. Robust allodynia was observed in all three ligation groups. Nerve degeneration confirmed that L4 and L5, respectively, are primary contributors to the tibial and sural branches of the sciatic nerve in mice. A larger von Frey sensitive area reflected the wider distribution of Wallerian degeneration in the hindlimb of L4- compared to L5-ligated mice. Ligation of mouse L4 and L5 spinal nerves produces consistent, robust neuropathic pain behaviors and is suitable as a model for investigating mechanisms of neuropathic pain and for testing of novel analgesics. Gabapentin, used as a validation drug in neuropathic pain models and as a reference compound for novel analgesics, significantly reduced allodynia in the mice tested (L4/L5 ligations). Given the ease of surgery, robust allodynia, and larger von Frey sensitive area, we conclude that combined ligation of spinal nerves L4 and L5 optimizes the SNL model in mice.

  18. Cell proliferation and apoptosis in optic nerve and brain integration centers of adult troutOncorhynchus mykiss after optic nerve injury

    Institute of Scientific and Technical Information of China (English)

    Evgeniya V Pushchina; Sachin Shukla; Anatoly A Varaksin; Dmitry K Obukhov

    2016-01-01

    Fishes have remarkable ability to effectively rebuild the structure of nerve cells and nerve ifbers after central nervous system injury. However, the underlying mechanism is poorly understood. In order to address this issue, we investigated the proliferation and apoptosis of cells in contralateral and ipsilateral optic nerves, after stab wound injury to the eye of an adult troutOncorhynchus mykiss. Heterogenous population of proliferating cells was investigated at 1 week after injury. TUNEL labeling gave a qualitative and quantita-tive assessment of apoptosis in the cells of optic nerve of trout 2 days after injury. After optic nerve injury, apoptotic response was investigated, and mass patterns of cell migration were found. The maximal con-centration of apoptotic bodies was detected in the areas of mass clumps of cells. It is probably indicative of massive cell death in the area of high phagocytic activity of macrophages/microglia. At 1 week after optic nerve injury, we observed nerve cell proliferation in the trout brain integration centers: the cerebellum and the optic tectum. In the optic tectum, proliferating cell nuclear antigen (PCNA)-immunopositive radial glia-like cells were identified. Proliferative activity of nerve cells was detected in the dorsal proliferative (matrix) area of the cerebellum and in parenchymal cells of the molecular and granular layers whereas local clusters of undifferentiated cells which formed neurogenic niches were observed in both the optic tectum and cerebellum after optic nerve injury.In vitro analysis of brain cells of trout showed that suspension cells compared with monolayer cells retain higher proliferative activity, as evidenced by PCNA immunolabeling. Phase contrast observation showed mitosis in individual cells and the formation of neurospheres which gradually increased during 1–4 days of culture. The present ifndings suggest that trout can be used as a novel model for studying neuronal regeneration.

  19. Differential presynaptic control of the synaptic effectiveness of cutaneous afferents evidenced by effects produced by acute nerve section.

    Science.gov (United States)

    Rudomin, P; Jiménez, I; Chávez, D

    2013-05-15

    In the anaesthetized cat, the acute section of the saphenous (Saph) and/or the superficial peroneal (SP) nerves was found to produce a long-lasting increase of the field potentials generated in the dorsal horn by stimulation of the medial branch of the sural (mSU) nerve. This facilitation was associated with changes in the level of the tonic primary afferent depolarization (PAD) of the mSU intraspinal terminals. The mSU afferent fibres projecting into Rexed's laminae III-IV were subjected to a tonic PAD that was reduced by the acute section of the SP and/or the Saph nerves. The mSU afferents projecting deeper into the dorsal horn (Rexed's laminae V-VI) were instead subjected to a tonic PAD that was increased after Saph and SP acute nerve section. A differential control of the synaptic effectiveness of the low-threshold cutaneous afferents according to their sites of termination within the dorsal horn is envisaged as a mechanism that allows selective processing of sensory information in response to tactile and nociceptive stimulation or during the execution of different motor tasks.

  20. NF-κB Regulates B-Cell-Derived Nerve Growth Factor Expression

    Institute of Scientific and Technical Information of China (English)

    Klaus Heese; Noriko Inoue; Tohru Sawada

    2006-01-01

    In the mammalian brain, four neurotrophins have been identified: nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3) and neurotrophin-4/5 (NT-4/5). NGF exerts an important role in the development and functions of the central and peripheral nervous system. However, it has recently been documented that several types of immune cells, such as mast cells, lymphocytes, basophils and eosinophils, produce,store and release NGF. Accumulating preclinical and clinical data indicate that dysfunctions of NGF and the other neurotrophins may contribute to impaired immune responses and concentration of NGF frequently correlates with disease severity. Thus, the aim of this study was to elucidate the potential signaling mechanisms of cytokineneurotrophins interactions contributing to increased NGF levels. Our data show that the transcription factorNF-κB plays a pivotal role in regulating B-cell-derived NGF expression.

  1. Repair of peripheral nerve defects with chemically extracted acellular nerve allografts loaded with neurotrophic factors-transfected bone marrow mesenchymal stem cells

    Institute of Scientific and Technical Information of China (English)

    Yan-ru Zhang; Ka Ka; Ge-chen Zhang; Hui Zhang; Yan Shang; Guo-qiang Zhao; Wen-hua Huang

    2015-01-01

    Chemically extracted acellular nerve allografts loaded with brain-derived neurotrophic fac-tor-transfected or ciliary neurotrophic factor-transfected bone marrow mesenchymal stem cells have been shown to repair sciatic nerve injury better than chemically extracted acellular nerve allografts alone, or chemically extracted acellular nerve allografts loaded with bone marrow mesenchymal stem cells. We hypothesized that these allografts compounded with both brain-derived neurotrophic factor- and ciliary neurotrophic factor-transfected bone marrow mesenchymal stem cells may demonstrate even better effects in the repair of peripheral nerve injury. We cultured bone marrow mesenchymal stem cells expressing brain-derived neuro-trophic factor and/or ciliary neurotrophic factor and used them to treat sciatic nerve injury in rats. We observed an increase in sciatic functional index, triceps wet weight recovery rate, myelin thickness, number of myelinated nerve ifbers, amplitude of motor-evoked potentials and nerve conduction velocity, and a shortened latency of motor-evoked potentials when al-lografts loaded with both neurotrophic factors were used, compared with allografts loaded with just one factor. Thus, the combination of both brain-derived neurotrophic factor and cili-ary neurotrophic factor-transfected bone marrow mesenchymal stem cells can greatly improve nerve injury.

  2. Horseradish peroxidase dye tracing and embryonic statoacoustic ganglion cell transplantation in the rat auditory nerve trunk.

    Science.gov (United States)

    Palmgren, Björn; Jin, Zhe; Jiao, Yu; Kostyszyn, Beata; Olivius, Petri

    2011-03-04

    At present severe damage to hair cells and sensory neurons in the inner ear results in non-treatable auditory disorders. Cell implantation is a potential treatment for various neurological disorders and has already been used in clinical practice. In the inner ear, delivery of therapeutic substances including neurotrophic factors and stem cells provide strategies that in the future may ameliorate or restore hearing impairment. In order to describe a surgical auditory nerve trunk approach, in the present paper we injected the neuronal tracer horseradish peroxidase (HRP) into the central part of the nerve by an intra cranial approach. We further evaluated the applicability of the present approach by implanting statoacoustic ganglion (SAG) cells into the same location of the auditory nerve in normal hearing rats or animals deafened by application of β-bungarotoxin to the round window niche. The HRP results illustrate labeling in the cochlear nucleus in the brain stem as well as peripherally in the spiral ganglion neurons in the cochlea. The transplanted SAGs were observed within the auditory nerve trunk but no more peripheral than the CNS-PNS transitional zone. Interestingly, the auditory nerve injection did not impair auditory function, as evidenced by the auditory brainstem response. The present findings illustrate that an auditory nerve trunk approach may well access the entire auditory nerve and does not compromise auditory function. We suggest that such an approach might compose a suitable route for cell transplantation into this sensory cranial nerve.

  3. [Construction of recombinant human nerve growth factor (rh-β-NGF) eukaryotic vector and its expression in HEK293 cells].

    Science.gov (United States)

    Li, Jingchuan; Xue, Bofu; Yuan, Yuan; Ma, Mo; Zhu, Lin; Milburn, Rebecca; Le, Li; Hu, Peizhen; Ye, Jing

    2015-03-01

    Human nerve growth factor (NGF) is a nerve cell growth regulation factor, which can provide nutrition for the neurons and promote the neurites outgrowth. In order to produce large-scale recombinant human nerve growth factor (rh-beta-NGF), we constructed a plasmid vector, which can stably express the rh-beta-NGF in the HEK293 cell lines. First, the plasmid of pCMV-beta-NGF-IRES-dhfr was constructed and transformed into HEK293 cells. Then MTX pressurized filter and limiting dilution methods were used to obtain monoclonal HEK293 cell lines. After stepwise reducing serum in culture media, the cells eventually adapted to serum-free medium and secreted rh-beta-NGF. SDS-PAGE analysis revealed that the expression product owned a molecular weight of about 13 kDa and a purity of more than 50%. The peptide mapping sequencing analysis demonstrated the sequences of rh-beta-NGF matched with the theoretical ones. Later we purified this protein by ion exchange and molecular sieve chromatograph. Finally, our experimental results exhibited that the recombinant cell lines can stably express rh-beta-NGF with a high efficiency of more than 20 pg/cell x day. In addition, this protein could successfully induce differentiation of PC12 cells. In summary, our recombinant HEK293 cells can express bio-active rh-beta-NGF with great efficiency and stability, which supply a valid basis to large-scale production of rh-beta-NGF.

  4. Methamphetamine-induced dopaminergic neurotoxicity and production of peroxynitrite are potentiated in nerve growth factor differentiated pheochromocytoma 12 cells.

    Science.gov (United States)

    Imam, Syed Z; Newport, Glenn D; Duhart, Helen M; Islam, Fakhrul; Slikker, William; Ali, Syed F

    2002-06-01

    Methamphetamine (METH) is a widely abused psychomotor stimulant known to cause dopaminergic neurotoxicity in rodents, nonhuman primates, and humans. METH administration selectively damages the dopaminergic nerve terminals, which is hypothesized to be due to release of dopamine from synaptic vesicles within the terminals. This process is believed to be mediated by the production of free radicals. The current study evaluates METH-induced dopaminergic toxicity in pheochromocytoma 12 (PC12) cells cultured in the presence or absence of nerve growth factor (NGF). Dopaminergic changes and the formation of 3-nitrotyrosine (3-NT), a marker for peroxynitrite production, were studied in PC12 cell cultures grown in the presence or absence of NGF after different doses of METH (100-1,000 microM). METH exposure did not cause significant alterations in cell viability and did not produce significant dopaminergic changes or 3-NT production in PC12 cells grown in NGF-negative media after 24 hours. However, cell viability of PC12 cells grown in NGF-positive media was decreased by 45%, and significant dose-dependent dopaminergic alteration and 3-NT production were observed 24 hours after exposure to METH. The current study supports the hypothesis that METH acts at the dopaminergic nerve terminals and produces dopaminergic damage by the production of free radical peroxynitrite.

  5. Transplantation of neural progenitor cells differentiated from adipose tissue-derived stem cells for treatment of sciatic nerve injury

    Institute of Scientific and Technical Information of China (English)

    Shasha Dong§; Na Liu§; Yang Hu ; Ping Zhang; Chao Pan; Youping Zhang; Yingxin Tang; Zhouping Tang 

    2016-01-01

    Objectives: Currently, the clinical repair of sciatic nerve injury remains difficult. Previous studies have confirmed that transplantation of adipose tissue-derived stem cells promotes nerve regeneration and restoration at peripheral nerve injury sites. Methods:In this study, adipose tissue-derived stem cells were induced to differentiate into neural progenitor cells, transfected with a green fluorescent protein-containing lentivirus, and then transplanted into the lesions of rats with sciatic nerve compression injury. Results: Fluorescence microscopy revealed that the transplanted cells survived, migrated, and differentiated in rats. At two weeks post-operation, a large number of transplanted cells had migrated to the injured lesions; at six weeks post-operation, transplanted cells were visible around the injured nerve and several cells were observed to express a Schwann cell marker. Sciatic function index and electrophysiological outcomes of the transplantation group were better than those of the control group. Cell transplantation promoted the recovery of motor nerve conduction velocity and com-pound muscle action potential amplitude, and reduced gastrocnemius muscle atrophy. Conclusions: Our experimental findings indicate that neural progenitor cells, differentiated from adipose tissue-derived stem cells, are potential seed stem cells that can be transplanted into lesions to treat sciatic nerve injury. This provides a theoretical basis for their use in clinical applications.

  6. Cancer stem cell-like cells derived from malignant peripheral nerve sheath tumors.

    Directory of Open Access Journals (Sweden)

    Melanie Spyra

    Full Text Available This study aims to examine whether or not cancer stem cells exist in malignant peripheral nerve sheath tumors (MPNST. Cells of established lines, primary cultures and freshly dissected tumors were cultured in serum free conditions supplemented with epidermal and fibroblast growth factors. From one established human MPNST cell line, S462, cells meeting the criteria for cancer stem cells were isolated. Clonal spheres were obtained, which could be passaged multiple times. Enrichment of stem cell-like cells in these spheres was also supported by increased expression of stem cell markers such as CD133, Oct4, Nestin and NGFR, and decreased expression of mature cell markers such as CD90 and NCAM. Furthermore, cells of these clonal S462 spheres differentiated into Schwann cells, smooth muscle/fibroblast and neurons-like cells under specific differentiation-inducing cultural conditions. Finally, subcutaneous injection of the spheres into immunodeficient nude mice led to tumor formation at a higher rate compared to the parental adherent cells (66% versus 10% at 2.5 × 10(5. These results provide evidence for the existence of cancer stem cell-like cells in malignant peripheral nerve sheath tumors.

  7. Nanoparticles carrying neurotrophin-3-modified Schwann cells promote repair of sciatic nerve defects

    Institute of Scientific and Technical Information of China (English)

    Haibin Zong; Hongxing Zhao; Yilei Zhao; Jingling Jia; Libin Yang; Chao Ma; Yang Zhang; Yuzhen Dong

    2013-01-01

    Schwann cells and neurotrophin-3 play an important role in neural regeneration, but the secretion of neurotrophin-3 from Schwann cells is limited, and exogenous neurotrophin-3 is inactived easily in vivo. In this study, we have transfected neurotrophin-3 into Schwann cells cultured in vitro using nanoparticle liposomes. Results showed that neurotrophin-3 was successfully transfected into Schwann cells, where it was expressed effectively and steadily. A composite of Schwann cells transfected with neurotrophin-3 and poly(lactic-co-glycolic acid) biodegradable conduits was transplanted into rats to repair 10-mm sciatic nerve defects. Transplantation of the composite scaffold could restore the myoelectricity and wave amplitude of the sciatic nerve by electrophysiological examination, promote nerve axonal and myelin regeneration, and delay apoptosis of spinal motor neurons. Experimental findings indicate that neurotrophin-3 transfected Schwann cells combined with bridge grafting can promote neural regeneration and functional recovery after nerve injury.

  8. Acceleration of Regeneration of Large-Gap Peripheral Nerve Injuries Using Acellular Nerve Allografts Plus Amniotic Fluid Derived Stem Cells (AFS)

    Science.gov (United States)

    2014-09-01

    nerves are incorporated into other rat strains without immunologic disturbances according to the vendor (AxoGen). 15. SUBJECT TERMS 16. SECURITY...mm). The outcomes of these surgeries will be compared to those obtained with autograft nerve repairs that currently have the best outcomes for large...ANA construct with AFS cells (Group 2), or with an autograft (nerve segment is cut out, reversed, and sewn back in place)(Group 3). All surgeries

  9. G-CSF prevents caspase 3 activation in Schwann cells after sciatic nerve transection, but does not improve nerve regeneration.

    Science.gov (United States)

    Frost, Hanna K; Kodama, Akira; Ekström, Per; Dahlin, Lars B

    2016-10-15

    Exogenous granulocyte-colony stimulating factor (G-CSF) has emerged as a drug candidate for improving the outcome after peripheral nerve injuries. We raised the question if exogenous G-CSF can improve nerve regeneration following a clinically relevant model - nerve transection and repair - in healthy and diabetic rats. In short-term experiments, distance of axonal regeneration and extent of injury-induced Schwann cell death was quantified by staining for neurofilaments and cleaved caspase 3, respectively, seven days after repair. There was no difference in axonal outgrowth between G-CSF-treated and non-treated rats, regardless if healthy Wistar or diabetic Goto-Kakizaki (GK) rats were examined. However, G-CSF treatment caused a significant 13% decrease of cleaved caspase 3-positive Schwann cells at the lesion site in healthy rats, but only a trend in diabetic rats. In the distal nerve segments of healthy rats a similar trend was observed. In long-term experiments of healthy rats, regeneration outcome was evaluated at 90days after repair by presence of neurofilaments, wet weight of gastrocnemius muscle, and perception of touch (von Frey monofilament testing weekly). The presence of neurofilaments distal to the suture line was similar in G-CSF-treated and non-treated rats. The weight ratio of ipsi-over contralateral gastrocnemius muscles, and perception of touch at any time point, were likewise not affected by G-CSF treatment. In addition, the inflammatory response in short- and long-term experiments was studied by analyzing ED1 stainable macrophages in healthy rats, but in neither case was any attenuation seen at the injury site or distal to it. G-CSF can prevent caspase 3 activation in Schwann cells in the short-term, but does not detectably affect the inflammatory response, nor improve early or late axonal outgrowth or functional recovery.

  10. Novel Method To Differentiate Human Embryonic Stem Cells Into Dopaminergic Nerve Cells | NCI Technology Transfer Center | TTC

    Science.gov (United States)

    The National Institute on Drug Abuse's Development and Plasticity Section is seeking statements of capability or interest from parties interested in collaborative research to further develop, evaluate, or commercialize novel methods to differentiate human embryonic stem cells into dopaminergic nerve cells. The invention described here is a novel method of differentiating human embryonic stem cells (hESCs) into dopaminergic nerve cells, which is preferable to the currently available dopaminergic differentiation techniques.

  11. Exposure to Nerve Growth Factor Worsens Nephrotoxic Effect Induced by Cyclosporine A in HK-2 Cells

    Science.gov (United States)

    Lofaro, Danilo; Toteda, Giuseppina; Lupinacci, Simona; Leone, Francesca; Gigliotti, Paolo; Papalia, Teresa; Bonofiglio, Renzo

    2013-01-01

    Nerve growth factor is a neurotrophin that promotes cell growth, differentiation, survival and death through two different receptors: TrkANTR and p75NTR. Nerve growth factor serum concentrations increase during many inflammatory and autoimmune diseases, glomerulonephritis, chronic kidney disease, end-stage renal disease and, particularly, in renal transplant. Considering that nerve growth factor exerts beneficial effects in the treatment of major central and peripheral neurodegenerative diseases, skin and corneal ulcers, we asked whether nerve growth factor could also exert a role in Cyclosporine A-induced graft nephrotoxicity. Our hypothesis was raised from basic evidence indicating that Cyclosporine A-inhibition of calcineurin-NFAT pathway increases nerve growth factor expression levels. Therefore, we investigated the involvement of nerve growth factor and its receptors in the damage exerted by Cyclosporine A in tubular renal cells, HK-2. Our results showed that in HK-2 cells combined treatment with Cyclosporine A + nerve growth factor induced a significant reduction in cell vitality concomitant with a down-regulation of Cyclin D1 and up-regulation of p21 levels respect to cells treated with Cyclosporine A alone. Moreover functional experiments showed that the co-treatment significantly up-regulated human p21promoter activity by involvement of the Sp1 transcription factor, whose nuclear content was negatively regulated by activated NFATc1. In addition we observed that the combined exposure to Cyclosporine A + nerve growth factor promoted an up-regulation of p75 NTR and its target genes, p53 and BAD leading to the activation of intrinsic apoptosis. Finally, the chemical inhibition of p75NTR down-regulated the intrinsic apoptotic signal. We describe two new mechanisms by which nerve growth factor promotes growth arrest and apoptosis in tubular renal cells exposed to Cyclosporine A. PMID:24244623

  12. Exposure to nerve growth factor worsens nephrotoxic effect induced by Cyclosporine A in HK-2 cells.

    Directory of Open Access Journals (Sweden)

    Donatella Vizza

    Full Text Available Nerve growth factor is a neurotrophin that promotes cell growth, differentiation, survival and death through two different receptors: TrkA(NTR and p75(NTR. Nerve growth factor serum concentrations increase during many inflammatory and autoimmune diseases, glomerulonephritis, chronic kidney disease, end-stage renal disease and, particularly, in renal transplant. Considering that nerve growth factor exerts beneficial effects in the treatment of major central and peripheral neurodegenerative diseases, skin and corneal ulcers, we asked whether nerve growth factor could also exert a role in Cyclosporine A-induced graft nephrotoxicity. Our hypothesis was raised from basic evidence indicating that Cyclosporine A-inhibition of calcineurin-NFAT pathway increases nerve growth factor expression levels. Therefore, we investigated the involvement of nerve growth factor and its receptors in the damage exerted by Cyclosporine A in tubular renal cells, HK-2. Our results showed that in HK-2 cells combined treatment with Cyclosporine A + nerve growth factor induced a significant reduction in cell vitality concomitant with a down-regulation of Cyclin D1 and up-regulation of p21 levels respect to cells treated with Cyclosporine A alone. Moreover functional experiments showed that the co-treatment significantly up-regulated human p21promoter activity by involvement of the Sp1 transcription factor, whose nuclear content was negatively regulated by activated NFATc1. In addition we observed that the combined exposure to Cyclosporine A + nerve growth factor promoted an up-regulation of p75 (NTR and its target genes, p53 and BAD leading to the activation of intrinsic apoptosis. Finally, the chemical inhibition of p75(NTR down-regulated the intrinsic apoptotic signal. We describe two new mechanisms by which nerve growth factor promotes growth arrest and apoptosis in tubular renal cells exposed to Cyclosporine A.

  13. Trigeminal nerve involvement in T-cell acute lymphoblastic leukemia: value of MR imaging

    Energy Technology Data Exchange (ETDEWEB)

    Karadag, Demet; Karaguelle, Ayse Tuba; Erden, Ilhan; Erden, Ayse E-mail: erden@ada.net.tr

    2002-10-01

    A 30-year-old male with T-cell acute lymphoblastic leukemia presented with facial numbness. Neurological examination revealed paresthesia of the left trigeminal nerve. Cerebrospinal fluid (CSF) cytology showed no atypical cells. Gadolinium-enhanced magnetic resonance (MR) imaging demonstrated enlargement and enhancement of intracranial portions of the left trigeminal nerve. The abnormal MR imaging findings almost completely resolved after the chemotherapy. Gadolinium-enhanced MR imaging is not only a useful procedure for the early diagnosis of cranial nerve invasion by leukemia but it might be helpful to follow the changes after the treatment.

  14. Schwann cells originating from skin-derived precursors promote peripheral nerve regeneration in rats

    Institute of Scientific and Technical Information of China (English)

    Ping Zhang; Xiaocheng Lu; Jianghai Chen; Zhenbing Chen

    2014-01-01

    Artiifcial guidance channels containing Schwann cells can promote the regeneration of injured peripheral nerve over long distances. However, primary Schwann cells are not suitable for autotransplantation. Under speciifc conditions, skin-derived progenitors can be induced to dif-ferentiate into Schwann cells. Therefore, adult rat dorsal skin (dermis)-derived progenitors were isolated and induced to differentiate with DMEM/F12 containing B27, neuregulin 1, and for-skolin. Immunolfuorescence staining and reverse transcription polymerase chain reaction (RT-PCR) conifrmed that the resultant cells were indeed Schwann cells. Artiifcial guidance channels containing skin-derived progenitors, Schwann cells originating from skin-derived progenitors, or primary Schwann cells were used to bridge 5 mm sciatic nerve defects. Schwann cells originating from skin-derived progenitors signiifcantly promoted sciatic nerve axonal regeneration. The sig-niifcant recovery of injured rat sciatic nerve function after the transplantation of Schwann cells originating from skin-derived progenitors was conifrmed by electromyogram. The therapeutic effect of Schwann cells originating from skin-derived progenitors was better than that of skin-de-rived progenitors. These findings indicate that Schwann cells originating from skin-derived precursors can promote peripheral nerve regeneration in rats.

  15. Extracellular stimulation of nerve cells with electric current spikes induced by voltage steps

    OpenAIRE

    2016-01-01

    A new stimulation paradigm is presented for the stimulation of nerve cells by extracellular electric currents. In the new paradigm stimulation is achieved with the current spike induced by a voltage step whenever the voltage step is applied to a live biological tissue. By experimental evidence and theoretical arguments, it is shown that this spike is well suited for the stimulation of nerve cells. Stimulation of the human tongue is used for proof of principle. Charge injection thresholds are ...

  16. A novel cell subset:Interferon-producing killer dendritic cells

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Recent reports introduce a novel cell subset of DCs with antigenic phenotypes shared by both NK cells and B cells, but without surface markers of pDCs and T cells, appearing to be a chimera of NK cells and DCs, namely interferon-producing killer dendritic cells(IKDCs).IKDCs not only secret type I and type II interferons to recognize and kill tumor cells effectively, but also express MHC-II molecules to present antigens.Thus, IKDCs are considered as important immunosurveilance cells for tumors, providing a link between innate and adaptive immunity.

  17. Electroactive biodegradable polyurethane significantly enhanced Schwann cells myelin gene expression and neurotrophin secretion for peripheral nerve tissue engineering.

    Science.gov (United States)

    Wu, Yaobin; Wang, Ling; Guo, Baolin; Shao, Yongpin; Ma, Peter X

    2016-05-01

    Myelination of Schwann cells (SCs) is critical for the success of peripheral nerve regeneration, and biomaterials that can promote SCs' neurotrophin secretion as scaffolds are beneficial for nerve repair. Here we present a biomaterials-approach, specifically, a highly tunable conductive biodegradable flexible polyurethane by polycondensation of poly(glycerol sebacate) and aniline pentamer, to significantly enhance SCs' myelin gene expression and neurotrophin secretion for peripheral nerve tissue engineering. SCs are cultured on these conductive polymer films, and the biocompatibility of these films and their ability to enhance myelin gene expressions and sustained neurotrophin secretion are successfully demonstrated. The mechanism of SCs' neurotrophin secretion on conductive films is demonstrated by investigating the relationship between intracellular Ca(2+) level and SCs' myelination. Furthermore, the neurite growth and elongation of PC12 cells are induced by adding the neurotrophin medium suspension produced from SCs-laden conductive films. These data suggest that these conductive degradable polyurethanes that enhance SCs' myelin gene expressions and sustained neurotrophin secretion perform great potential for nerve regeneration applications.

  18. Stem Cell Transplantation for Peripheral Nerve Regeneration: Current Options and Opportunities

    Directory of Open Access Journals (Sweden)

    Liangfu Jiang

    2017-01-01

    Full Text Available Peripheral nerve regeneration is a complicated process highlighted by Wallerian degeneration, axonal sprouting, and remyelination. Schwann cells play an integral role in multiple facets of nerve regeneration but obtaining Schwann cells for cell-based therapy is limited by the invasive nature of harvesting and donor site morbidity. Stem cell transplantation for peripheral nerve regeneration offers an alternative cell-based therapy with several regenerative benefits. Stem cells have the potential to differentiate into Schwann-like cells that recruit macrophages for removal of cellular debris. They also can secrete neurotrophic factors to promote axonal growth, and remyelination. Currently, various types of stem cell sources are being investigated for their application to peripheral nerve regeneration. This review highlights studies involving the stem cell types, the mechanisms of their action, methods of delivery to the injury site, and relevant pre-clinical or clinical data. The purpose of this article is to review the current point of view on the application of stem cell based strategy for peripheral nerve regeneration.

  19. Stem cell therapy for central nerve system injuries:glial cells hold the key

    Institute of Scientific and Technical Information of China (English)

    Li Xiao; Chikako Saiki; Ryoji Ide

    2014-01-01

    Mammalian adult central nerve system (CNS) injuries are devastating because of the intrinsic dififculties for effective neuronal regeneration. The greatest problem to be overcome for CNS recovery is the poor regeneration of neurons and myelin-forming cells, oligodendrocytes. En-dogenous neural progenitors and transplanted exogenous neuronal stem cells can be the source for neuronal regeneration. However, because of the harsh local microenvironment, they usually have very low efifcacy for functional neural regeneration which cannot compensate for the loss of neurons and oligodendrocytes. Glial cells (including astrocytes, microglia, oligodendrocytes and NG2 glia) are the majority of cells in CNS that provide support and protection for neurons. Inside the local microenvironment, glial cells largely inlfuence local and transplanted neural stem cells survival and fates. This review critically analyzes current ifnding of the roles of glial cells in CNS regeneration, and highlights strategies for regulating glial cells’ behavior to create a permis-sive microenvironment for neuronal stem cells.

  20. Dark rearing maintains tyrosine hydroxylase expression in retinal amacrine cells following optic nerve transection

    Institute of Scientific and Technical Information of China (English)

    Wei Wan; Zhenghai Liu; Xiaosheng Wang; Xuegang Luo

    2012-01-01

    The present study examined changes in retinal tyrosine hydroxylase (TH) expression in rats having undergone optic nerve transection and housed under a normal day/night cycle or in the dark. The aim was to investigate the effects of amacrine cells on axonal regeneration in retinal ganglion cells and on the synapses that transmit visual signals. The results revealed that retinal TH expression gradually decreased following optic nerve transection in rats housed under a normal day/night cycle, reaching a minimum at 5 days. In contrast, retinal TH expression decreased to a minimum at 1 day following optic nerve transection in dark reared rats, gradually increasing afterward and reaching a normal level at 5-7 days. The number of TH-positive synaptic particles correlated with the TH levels, indicating that dark rearing can help maintain TH expression during the synaptic degeneration stage (5-7 days after optic nerve injury) in retinal amacrine cells.

  1. Macrophage polarization in nerve injury:do Schwann cells play a role?

    Institute of Scientific and Technical Information of China (English)

    Jo Anne Stratton; Prajay T Shah

    2016-01-01

    In response to peripheral nerve injury, the inlfammatory response is almost entirely comprised of inifltrat-ing macrophages. Macrophages are a highly plastic, heterogenic immune cell, playing an indispensable role in peripheral nerve injury, clearing debris and regulating the microenvironment to allow for efifcient regen-eration. There are several cells within the microenvironment that likely interact with macrophages to support their function –most notably the Schwann cell, the glial cell of the peripheral nervous system. Schwann cells express several ligands that are known to interact with receptors expressed by macrophages, yet the effects of Schwann cells in regulating macrophage phenotype remains largely unexplored. This review discusses macrophages in peripheral nerve injury and how Schwann cells may regulate their behavior.

  2. Senescence in adipose-derived stem cells and its implications in nerve regeneration

    Institute of Scientific and Technical Information of China (English)

    Cristina Mantovani; Giorgio Terenghi; Valerio Magnaghi

    2014-01-01

    Adult mesenchymal stem cells, specifically adipose-derived stem cells have self-renewal and multiple differentiation potentials and have shown to be the ideal candidate for therapeutic applications in regenerative medicine, particularly in peripheral nerve regeneration. Adipose-de-rived stem cells are easily harvested, although they may show the effects of aging, hence their potential in nerve repair may be limited by cellular senescence or donor age. Cellular senescence is a complex process whereby stem cells grow old as consequence of intrinsic events (e.g., DNA damage) or environmental cues (e.g., stressful stimuli or diseases), which determine a permanent growth arrest. Several mechanisms are implicated in stem cell senescence, although no one is exclusive of the others. In this review we report some of the most important factors modulating the senescence process, which can inlfuence adipose-derived stem cell morphology and function, and compromise their clinical application for peripheral nerve regenerative cell therapy.

  3. Platelet-rich plasma gel in combination with Schwann cells for repair of sciatic nerve injury

    Institute of Scientific and Technical Information of China (English)

    Fagang Ye; Haiyan Li; Guangxi Qiao; Feng Chen; Hao Tao; Aiyu Ji; Yanling Hu

    2012-01-01

    Bone marrow mesenchymal stem cells were isolated from New Zealand white rabbits,culture-expanded and differentiated into Schwann cell-like cells.Autologous platelet-rich plasma and Schwann cell-like cells were mixed in suspension at a density of 1 × 106 cells/mL,prior to introduction into a poly (lactic-co-glycolic acid) conduit.Fabricated tissue-engineered nerves were implanted into rabbits to bridge 10 mm sciatic nerve defects (platelet-rich plasma group).Controls were established using fibrin as the seeding matrix for Schwann cell-like cells at identical density to construct tissue-engineered nerves (fibrin group).Twelve weeks after implantation,toluidine blue staining and scanning electron microscopy were used to demonstrate an increase in the number of regenerating nerve fibers and thickness of the myelin sheath in the platelet-rich plasma group compared with the fibrin group.Fluoro-gold retrograde labeling revealed that the number of Fluoro-gold-positive neurons in the dorsal root ganglion and the spinal cord anterior horn was greater in the platelet-rich plasma group than in the fibrin group.Electrophysiological examination confirmed that compound muscle action potential and nerve conduction velocity were superior in the platelet-rich plasma group compared with the fibrin group.These results indicate that autologous platelet-rich plasma gel can effectively serve as a seeding matrix for Schwann cell-like cells to construct tissue-engineered nerves to promote peripheral nerve regeneration.

  4. Nerve growth factor promotes in vitro proliferation of neural stem cells from tree shrews

    Directory of Open Access Journals (Sweden)

    Liu-lin Xiong

    2016-01-01

    Full Text Available Neural stem cells promote neuronal regeneration and repair of brain tissue after injury, but have limited resources and proliferative ability in vivo. We hypothesized that nerve growth factor would promote in vitro proliferation of neural stem cells derived from the tree shrews, a primate-like mammal that has been proposed as an alternative to primates in biomedical translational research. We cultured neural stem cells from the hippocampus of tree shrews at embryonic day 38, and added nerve growth factor (100 µg/L to the culture medium. Neural stem cells from the hippocampus of tree shrews cultured without nerve growth factor were used as controls. After 3 days, fluorescence microscopy after DAPI and nestin staining revealed that the number of neurospheres and DAPI/nestin-positive cells was markedly greater in the nerve growth factor-treated cells than in control cells. These findings demonstrate that nerve growth factor promotes the proliferation of neural stem cells derived from tree shrews.

  5. Distribution of Neuron Cell Bodies in the Intraspinal Portion of the Spinal Accessory Nerve in Humans.

    Science.gov (United States)

    Boehm, Karl E; Kondrashov, Peter

    2016-01-01

    The spinal accessory nerve is often identified as a purely motor nerve innervating the trapezius and sternocleidomastoid muscles. Although it may contain proprioceptive neurons found in cervical spinal levels C2-C4, limited research has focused on the histology of the spinal accessory nerve. The objective of the present study was to examine the spinal accessory nerve to determine if there are neuronal cell bodies within the spinal accessory nerve in humans. Cervical spinal cords were dissected from eight cadavers that had previously been used for dissection in other body regions. The segmental rootlets were removed to quantify the neuron cell bodies present at each spinal level. Samples were embedded in paraffin; sectioned; stained with hematoxylin and eosin; and examined using a microscope at 4×, 10×, and 40× magnification. Digital photography was used to image the samples. Neuronal cell bodies were found in 100% of the specimens examined, with non-grossly visible ganglia found at spinal levels C1-C4. The C1 spinal level of the spinal accessory nerve had the highest number of neuron cell bodies.

  6. Schwann cells express erythropoietin receptor and represent a major target for Epo in peripheral nerve injury.

    Science.gov (United States)

    Li, Xiaoqing; Gonias, Steven L; Campana, W Marie

    2005-09-01

    Erythropoietin (Epo) expresses potent neuroprotective activity in the peripheral nervous system; however, the underlying mechanism remains incompletely understood. In this study, we demonstrate that Epo is upregulated in sciatic nerve after chronic constriction injury (CCI) and crush injury in rats, largely due to local Schwann cell production. In uninjured and injured nerves, Schwann cells also express Epo receptor (EpoR), and its expression is increased during Wallerian degeneration. CCI increased the number of Schwann cells at the injury site and the number was further increased by exogenously administered recombinant human Epo (rhEpo). To explore the activity of Epo in Schwann cells, primary cultures were established. These cells expressed cell-surface Epo receptors, with masses of 71 and 62 kDa, as determined by surface protein biotinylation and affinity precipitation. The 71-kDa species was rapidly but transiently tyrosine-phosphorylated in response to rhEpo. ERK/MAP kinase was also activated in rhEpo-treated Schwann cells; this response was blocked by pharmacologic antagonism of JAK-2. RhEpo promoted Schwann cell proliferation, as determined by BrdU incorporation. Cell proliferation was ERK/MAP kinase-dependent. These results support a model in which Schwann cells are a major target for Epo in injured peripheral nerves, perhaps within the context of an autocrine signaling pathway. EpoR-induced cell signaling and Schwann cell proliferation may protect injured peripheral nerves and promote regeneration.

  7. Axonal and Schwann cell BACE1 is equally required for remyelination of peripheral nerves.

    Science.gov (United States)

    Hu, Xiangyou; Hu, Jinxuan; Dai, Lu; Trapp, Bruce; Yan, Riqiang

    2015-03-04

    Inhibition of β-site APP cleaving enzyme 1 (BACE1) is being pursued as a therapeutic target for treating patients with Alzheimer's disease because BACE1 is the sole β-secretase for generating β-amyloid peptide. Knowledge regarding the other cellular functions of BACE1 is therefore critical for the safe use of BACE1 inhibitors in human patients. BACE1 deficiency in mice causes hypomyelination during development and impairs remyelination in injured sciatic nerves. Since BACE1 is expected to be ubiquitously expressed, we asked whether axonal or Schwann cell BACE1 is required for optimal remyelination. By swapping sciatic nerve segments from BACE1-null mice with the corresponding wild-type nerve segments or vice versa, we tested how a deficiency of BACE1 in Schwann cells or axons affects remyelination. Our results show that BACE1 in axons and Schwann cells is similarly important for remyelination of regenerated axons. Nerve injury induces BACE1 transcription and protein levels are elevated in Schwann cells. Expression of type I neuregulin 1 (Nrg1), rather than type III Nrg1, was induced by Schwann cells, and the abolished Nrg1 cleavage in BACE1-null Schwann cells contributed to decreased remyelination of regenerated axons. Hence, this study is the first to demonstrate the equal importance of axonal and Schwann cell BACE1 for remyelination of injured nerves.

  8. [Changes in neuropeptide Y and substance P immunoreactive nerve fibres and immunocompetent cells in hepatitis].

    Science.gov (United States)

    Fehér, Erzsébet

    2015-11-22

    Neuropeptide Y and substance P were thought to play a role in the function of immune cells and in amplification or elimination of the inflammatory processes. In hepatitis the number of both neuropeptide Y and substance P immunoreactive nerve fibres are increased, where the increase of neoropeptide Y is significant. A large number of lymphocytes and mast cells are also stained for neuropeptide Y and substance P. Very close associations (less than 1 µm) were observed between neuropeptide Y immunoreactive nerve fibres and immune cells stained also with neuropeptide Y. Some immune cells were also found to be immunoreactive for tumor necrosis factor-α and NF-κB. Some of the SP IR immunocells were also stained for TNF-α and nuclear factor kappaB. Based on these data it is hypothesized that neuropeptid Y and substance P released from nerve fibres and immune cells play a role in inflammation and elimination of inflammation in hepatitis.

  9. Macrophage-Induced Blood Vessels Guide Schwann Cell-Mediated Regeneration of Peripheral Nerves.

    Science.gov (United States)

    Cattin, Anne-Laure; Burden, Jemima J; Van Emmenis, Lucie; Mackenzie, Francesca E; Hoving, Julian J A; Garcia Calavia, Noelia; Guo, Yanping; McLaughlin, Maeve; Rosenberg, Laura H; Quereda, Victor; Jamecna, Denisa; Napoli, Ilaria; Parrinello, Simona; Enver, Tariq; Ruhrberg, Christiana; Lloyd, Alison C

    2015-08-27

    The peripheral nervous system has remarkable regenerative capacities in that it can repair a fully cut nerve. This requires Schwann cells to migrate collectively to guide regrowing axons across a 'bridge' of new tissue, which forms to reconnect a severed nerve. Here we show that blood vessels direct the migrating cords of Schwann cells. This multicellular process is initiated by hypoxia, selectively sensed by macrophages within the bridge, which via VEGF-A secretion induce a polarized vasculature that relieves the hypoxia. Schwann cells then use the blood vessels as "tracks" to cross the bridge taking regrowing axons with them. Importantly, disrupting the organization of the newly formed blood vessels in vivo, either by inhibiting the angiogenic signal or by re-orienting them, compromises Schwann cell directionality resulting in defective nerve repair. This study provides important insights into how the choreography of multiple cell-types is required for the regeneration of an adult tissue.

  10. Effect on pancreatic beta cells and nerve cells by low let x-ray

    Energy Technology Data Exchange (ETDEWEB)

    Park, Kwang Hun [Dept. of Nuclear Medicine, Kyungbuk National University Hospital, Daegu (Korea, Republic of); Kim, Kgu Hwan [Dept. of Radiological Technology, Daegu health College, Daegu (Korea, Republic of)

    2014-03-15

    Cultured pancreatic beta cells and nerve cells, it is given normal condition of 10% FBS (fetal bovine serum), 11.1 mM glucose and hyperglycemia condition of 1% FBS, 30 mM glucose. For low LET X-ray irradiated with 0.5 Gy/hr dose-rate(total dose: 0.5 to 5 Gy). Survival rates were measured by MTT assay. When non irradiated, differentiated in the pancreatic beta cells experiment is hyperglycemia conditions survival rate compared to normal conditions survival rate seemed a small reduction. However increasing the total dose of X-ray, the survival rate of normal conditions decreased slightly compared to the survival rate of hyperglycemia conditions, the synergistic effect was drastically reduced. When non irradiated, undifferentiated in the nerve cells experiment is hyperglycemia conditions survival rate compared to normal conditions survival rate seemed a large reduction. As the cumulative dose of X-ray normal conditions and hyperglycemia were all relatively rapid cell death. But the rate of decreased survivals by almost parallel to the reduction proceed and it didn't show synergistic effect.

  11. The neurotoxic effects of hydrogen peroxide and copper in Retzius nerve cells of the leech Haemopis sanguisuga

    Directory of Open Access Journals (Sweden)

    Zorica D. Jovanovic

    2016-04-01

    Full Text Available Oxidative stress and the generation of reactive oxygen species (ROS play an important role in cellular damage. Electrophysiological analyses have shown that membrane transport proteins are susceptible to ROS. In the present study, oxidative stress was induced in Retzius nerve cells of the leech Haemopis sanguisuga by bath application of 1 mM of hydrogen peroxide (H2O2 and 0.02 mM of copper (Cu for 20 min. The H2O2/Cu(II produced considerable changes in the electrical properties of the Retzius nerve cells. Intracellular recording of the resting membrane potential revealed that the neuronal membrane was depolarized in the presence of H2O2/Cu(II. We found that the amplitude of action potentials decreased, while the duration augmented in a progressive way along the drug exposure time. The combined application of H2O2 and Cu(II caused an initial excitation followed by depression of the spontaneous electrical activity. Voltage-clamp recordings revealed a second effect of the oxidant, a powerful inhibition of the outward potassium channels responsible for the repolarization of action potentials. The neurotoxic effect of H2O2/Cu(II on the spontaneous spike electrogenesis and outward K+ current of Retzius nerve cells was reduced in the presence of hydroxyl radical scavengers, dimethylthiourea and dimethyl sulfoxide, but not mannitol. This study provides evidence for the oxidative modification of outward potassium channels in Retzius nerve cells. The oxidative mechanism of the H2O2/Cu(II system action on the electrical properties of Retzius neurons proposed in this study might have a wider significance, referring not only to leeches but also to mammalian neurons.

  12. Engrafted human induced pluripotent stem cell-derived anterior specified neural progenitors protect the rat crushed optic nerve.

    Directory of Open Access Journals (Sweden)

    Leila Satarian

    Full Text Available BACKGROUND: Degeneration of retinal ganglion cells (RGCs is a common occurrence in several eye diseases. This study examined the functional improvement and protection of host RGCs in addition to the survival, integration and neuronal differentiation capabilities of anterior specified neural progenitors (NPs following intravitreal transplantation. METHODOLOGY/PRINCIPAL FINDINGS: NPs were produced under defined conditions from human induced pluripotent stem cells (hiPSCs and transplanted into rats whose optic nerves have been crushed (ONC. hiPSCs were induced to differentiate into anterior specified NPs by the use of Noggin and retinoic acid. The hiPSC-NPs were labeled by green fluorescent protein or a fluorescent tracer 1,1' -dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI and injected two days after induction of ONC in hooded rats. Functional analysis according to visual evoked potential recordings showed significant amplitude recovery in animals transplanted with hiPSC-NPs. Retrograde labeling by an intra-collicular DiI injection showed significantly higher numbers of RGCs and spared axons in ONC rats treated with hiPSC-NPs or their conditioned medium (CM. The analysis of CM of hiPSC-NPs showed the secretion of ciliary neurotrophic factor, basic fibroblast growth factor, and insulin-like growth factor. Optic nerve of cell transplanted groups also had increased GAP43 immunoreactivity and myelin staining by FluoroMyelin™ which imply for protection of axons and myelin. At 60 days post-transplantation hiPSC-NPs were integrated into the ganglion cell layer of the retina and expressed neuronal markers. CONCLUSIONS/SIGNIFICANCE: The transplantation of anterior specified NPs may improve optic nerve injury through neuroprotection and differentiation into neuronal lineages. These NPs possibly provide a promising new therapeutic approach for traumatic optic nerve injuries and loss of RGCs caused by other diseases.

  13. Mesenchymal stem cells modified with nerve growth factor improve recovery of the inferior alveolar nerve after mandibular distraction osteogenesis in rabbits.

    Science.gov (United States)

    Wang, L; Zhao, Y; Cao, J; Yang, X; Lei, D

    2015-03-01

    Distraction osteogenesis is widely used in the treatment of bony deformities and defects. However, injury to the inferior alveolar nerve is a concern. Our aim was to investigate the feasibility of using lentiviral-mediated human nerve growth factor beta (hNGFβ) of the inferior alveolar nerve in mandibular distraction osteogenesis in rabbits. To achieve this, mesenchymal stem cells (MSC) from the bone marrow of rabbit mandibles were isolated and genetically engineered using recombinant lentiviral vector containing hNGFβ. Twenty New Zealand white rabbits underwent mandibular distraction osteogenesis, and 5 million MSC transduced with hNGFβ-vector or control vector were transplanted around the nerve in the gap where the bone had been fractured during the operation (n=10 in each group). After gradual distraction, samples of the nerve were harvested for histological and histomorphometric analysis. We found that the genetically engineered MSC transduced by the lentiviral vector were able to secrete hNGFβ at physiologically relevant concentrations as measured by ELISA. Histological examination of the nerve showed more regenerating nerve fibres and less myelin debris in the group in which hNGFβ-modified MSC had been implanted than in the control group. Histomorphometric analysis of the nerve showed increased density of myelinated fibres in the group in which hNGFβ-modified MSC had been implanted than in the control group. The data suggest that implantation of hNGFβ-modified MSC can accelerate the morphological recovery of the inferior alveolar nerve during mandibular distraction osteogenesis in rabbits. The use of lentiviral-mediated gene treatment to deliver hNGFβ through MSC may be a promising way of minimising injury to the nerve.

  14. Biological conduits combining bone marrow mesenchymal stem cells and extracellular matrix to treat long-segment sciatic nerve defects

    Directory of Open Access Journals (Sweden)

    Yang Wang

    2015-01-01

    Full Text Available The transplantation of polylactic glycolic acid conduits combining bone marrow mesenchymal stem cells and extracellular matrix gel for the repair of sciatic nerve injury is effective in some respects, but few data comparing the biomechanical factors related to the sciatic nerve are available. In the present study, rabbit models of 10-mm sciatic nerve defects were prepared. The rabbit models were repaired with autologous nerve, a polylactic glycolic acid conduit + bone marrow mesenchymal stem cells, or a polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel. After 24 weeks, mechanical testing was performed to determine the stress relaxation and creep parameters. Following sciatic nerve injury, the magnitudes of the stress decrease and strain increase at 7,200 seconds were largest in the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel group, followed by the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells group, and then the autologous nerve group. Hematoxylin-eosin staining demonstrated that compared with the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells group and the autologous nerve group, a more complete sciatic nerve regeneration was found, including good myelination, regularly arranged nerve fibers, and a completely degraded and resorbed conduit, in the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel group. These results indicate that bridging 10-mm sciatic nerve defects with a polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel construct increases the stress relaxation under a constant strain, reducing anastomotic tension. Large elongations under a constant physiological load can limit the anastomotic opening and shift, which is beneficial for the regeneration and functional reconstruction of sciatic nerve. Better

  15. Olfactory ensheathing cell transplantation for a patient with chronic sciatic nerve injury

    Directory of Open Access Journals (Sweden)

    Zhang F

    2016-12-01

    Full Text Available Feng Zhang,1,2 Xiangzhi Meng,2 Fang Lu,2 Aixian Liu,2 Hongyun Huang1,2 1Cell Therapy Center, Beijing Hongtianji Neuroscience Academy, 2Neurorehabilitation Center, Beijing Rehabilitation Hospital, Capital Medical University, Beijing, People’s Republic of China Objective: To observe the result of olfactory ensheathing cell (OEC transplantation in a patient with chronic sciatic nerve injury. Case report: A 53-year-old male patient with chronic (1 year sciatic nerve injury on left side received OEC transplantation at the lesion site. He received follow-up assessment according to the American Spinal Injury Association standard at 10 days, 6 months, and 1 year after OEC therapy. The muscle strength of his left lower limb increased and numbness decreased during the early stage of cell therapy. His motor function improved with each evaluation. His limp walking gait recovered, and numbness sensation got nearly normal after 1 year of follow-up. There were no side effects. Conclusion: OEC transplantation may be an option for chronic peripheral (sciatic nerve injury. Keywords: olfactory ensheathing cell transplantation, sciatic nerve injury, peripheral nerve injury, function improvement, neurorestoration

  16. Intestinal epithelial stem/progenitor cells are controlled by mucosal afferent nerves.

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    Ove Lundgren

    Full Text Available BACKGROUND: The maintenance of the intestinal epithelium is of great importance for the survival of the organism. A possible nervous control of epithelial cell renewal was studied in rats and mice. METHODS: Mucosal afferent nerves were stimulated by exposing the intestinal mucosa to capsaicin (1.6 mM, which stimulates intestinal external axons. Epithelial cell renewal was investigated in the jejunum by measuring intestinal thymidine kinase (TK activity, intestinal (3H-thymidine incorporation into DNA, and the number of crypt cells labeled with BrdU. The influence of the external gut innervation was minimized by severing the periarterial nerves. PRINCIPAL FINDINGS: Luminal capsaicin increased all the studied variables, an effect nervously mediated to judge from inhibitory effects on TK activity or (3H-thymidine incorporation into DNA by exposing the mucosa to lidocaine (a local anesthetic or by giving four different neurotransmitter receptor antagonists i.v. (muscarinic, nicotinic, neurokinin1 (NK1 or calcitonin gene related peptide (CGRP receptors. After degeneration of the intestinal external nerves capsaicin did not increase TK activity, suggesting the involvement of an axon reflex. Intra-arterial infusion of Substance P (SP or CGRP increased intestinal TK activity, a response abolished by muscarinic receptor blockade. Immunohistochemistry suggested presence of M3 and M5 muscarinic receptors on the intestinal stem/progenitor cells. We propose that the stem/progenitor cells are controlled by cholinergic nerves, which, in turn, are influenced by mucosal afferent neuron(s releasing acetylcholine and/or SP and/or CGRP. In mice lacking the capsaicin receptor, thymidine incorporation into DNA and number of crypt cells labeled with BrdU was lower than in wild type animals suggesting that nerves are important also in the absence of luminal capsaicin, a conclusion also supported by the observation that atropine lowered thymidine incorporation into DNA

  17. An animal model of peripheral nerve regeneration after the application of a collagen-polyvinyl alcohol scaffold and mesenchymal stem cells.

    Science.gov (United States)

    Marinescu, Silviu Adrian; Zărnescu, Otilia; Mihai, Ioana Ruxandra; Giuglea, Carmen; Sinescu, Ruxandra Diana

    2014-01-01

    Extensive nerve injuries often leading to nerve gaps can benefit, besides the gold standard represented by autologous nerve grafts, by the inciting field of tissue engineering. To enhance the role of biomaterials in nerve regeneration, the nerve conduits are associated with Schwann or Schwann-like cells. In this study, we evaluated rat sciatic nerve regeneration, by using a biodegradable nerve guide composed of Collagen (COL) and Polyvinyl Alcohol (PVA), associated with mesenchymal stem cells (MSC). After the exposure of the rat sciatic nerve, a nerve gap was created by excising 1 cm of the nerve. Three experimental groups were used for nerve gap bridging: autografts, nerve conduits filled with medium culture and nerve conduits filled with MSC. The methods of sensory and motor assessment consisted of the functional evaluation of sciatic nerve recovery - toe-spread, pinprick tests and gastrocnemius muscle index (GMI). The histological and immunocytochemical analysis of the probes that were harvested from the repair site was performed at 12 weeks. Successful nerve regeneration was noted in all three groups at the end of the 12th week. The functional and immunocytochemical results suggested that COL-PVA tubes supported with mesenchymal stem cells could be considered similar to autologous nerve grafts in peripheral nerve regeneration, without the drawbacks of the last ones. The functional results were better for the autografts and the ultrastructural data were better for the nerve conduits, but there were not noticed any statistical differences.

  18. Lacrimal gland and perioptic nerve lesions due to Langerhans cell histiocytosis (2007: 9b)

    Energy Technology Data Exchange (ETDEWEB)

    Herman, M.; Demaerel, P.; Wilms, G. [University Hospitals Leuven, Department of Radiology, Leuven (Belgium); Gool, S. van [University Hospitals Leuven, Department of Pediactrics, Leuven (Belgium); Casteels, I. [University Hospitals Leuven, Department of Ophthalmology, Leuven (Belgium)

    2007-12-15

    We report a patient presenting with bilateral lacrimal gland involvement and perioptic nerve sheath lesions due to Langerhans cell histiocytosis (LCH) invasion. LCH is a rare multisystemic disease characterized by a clonal proliferation of Langerhans cells. All organs may be involved with a clinical spectrum ranging from a solitary bone lesion to a severe life-threatening multisystem disease. Osteolytic orbital bone lesions with extension into the adjacent orbital soft tissues have been described. To our knowledge, lacrimal gland involvement has probably been described only once before. Perioptic nerve lesions are also very rare, having been described only three times before. (orig.)

  19. Nerve growth factor modulate proliferation of cultured rabbit corneal endothelial cells and epithelial cells.

    Science.gov (United States)

    Li, Xinyu; Li, Zhongguo; Qiu, Liangxiu; Zhao, Changsong; Hu, Zhulin

    2005-01-01

    In order to investigate the effect of nerve growth factor (NGF) on the proliferation of rabbit corneal endothelial cells and epithelial cells, the in vitro cultured rabbit corneal endothelial cells and epithelial cells were treated with different concentrations of NGF. MTT assay was used to examine the clonal growth and proliferation of the cells by determining the absorbency values at 570 nm. The results showed that NGF with three concentrations ranging from 5 U/mL to 500 U/mL enhanced the proliferation of rabbit corneal endothelial cells in a concentration-dependent manner. 50 U/mL and 500 U/mL NGF got more increase of proliferation than that of 5 U/mL NGF did. Meanwhile, 50 U/mL and 500 U/mL NGF could promote the proliferation of the rabbit corneal epithelial cells significantly in a concentration-dependent manner. However, 5 U/mL NGF did not enhance the proliferation of epithelial cells. It was suggested that exogenous NGF can stimulate the proliferation of both rabbit corneal endothelial and epithelial cells, but the extent of modulation is different.

  20. Producing primate embryonic stem cells by somatic cell nuclear transfer.

    Science.gov (United States)

    Byrne, J A; Pedersen, D A; Clepper, L L; Nelson, M; Sanger, W G; Gokhale, S; Wolf, D P; Mitalipov, S M

    2007-11-22

    Derivation of embryonic stem (ES) cells genetically identical to a patient by somatic cell nuclear transfer (SCNT) holds the potential to cure or alleviate the symptoms of many degenerative diseases while circumventing concerns regarding rejection by the host immune system. However, the concept has only been achieved in the mouse, whereas inefficient reprogramming and poor embryonic development characterizes the results obtained in primates. Here, we used a modified SCNT approach to produce rhesus macaque blastocysts from adult skin fibroblasts, and successfully isolated two ES cell lines from these embryos. DNA analysis confirmed that nuclear DNA was identical to donor somatic cells and that mitochondrial DNA originated from oocytes. Both cell lines exhibited normal ES cell morphology, expressed key stem-cell markers, were transcriptionally similar to control ES cells and differentiated into multiple cell types in vitro and in vivo. Our results represent successful nuclear reprogramming of adult somatic cells into pluripotent ES cells and demonstrate proof-of-concept for therapeutic cloning in primates.

  1. Human umbilical cord Wharton's Jelly-derived mesenchymal stem cells differentiation into nerve-like cells

    Institute of Scientific and Technical Information of China (English)

    MA Lian; FENG Xue-yong; CUI Bing-lin; Frieda Law; JIANG Xue-wu; YANG Li-ye; XIE Qing-dong; HUANG Tian-hua

    2005-01-01

    Background The two most basic properties of mesenchymal stem cells (MSCs) are the capacities to self-renew indefinitely and differentiate into multiple cells and tissue types. The cells from human umbilical cord Wharton's Jelly have properties of MSCs and represent a rich source of primitive cells. This study was conducted to explore the possibility of inducing human umbilical cord Wharton's Jelly-derived MSCs to differentiate into nerve-like cells.Methods MSCs were cultured from the Wharton's Jelly taken from human umbilical cord of babies delivered after full-term normal labor. Salvia miltiorrhiza and β-mercaptoethanol were used to induce the human umbilical cord-derived MSCs to differentiate. The expression of neural protein markers was shown by immunocytochemistry. The induction process was monitored by phase contrast microscopy, electron microscopy (EM), and laser scanning confocal microscopy (LSCM) .The pleiotrophin and nestin genes were measured by reverse transcription-polymerase chain reaction (RT-PCR). Results MSCs in the Wharton's Jelly were easily attainable and could be maintained and expanded in culture. They were positive for markers of MSCs, but negative for markers of hematopoietic cells and graft-versus-host disease (GVHD)-related cells. Treatment with Salvia miltiorrhiza caused Wharton's Jelly cells to undergo profound morphological changes. The induced MSCs developed rounded cell bodies with multiple neurite-like extensions. Eventually they developed processes that formed networks reminiscent of primary cultures of neurons. Salvia miltiorrhiza and β-mercaptoethanol also induced MSCs to express nestin, β-tubulinⅢ, neurofilament (NF) and glial fibrillary acidic protein (GFAP). It was confirmed by RT-PCR that MSCs could express pleiotrophin both before and after induction by Salvia miltiorrhiza. The expression was markedly enhanced after induction and the nestin gene was also expressed.Conclusions MSCs could be isolated from human umbilical

  2. Human umbilical cord mesenchymal stem cells promote peripheral nerve repairvia paracrine mechanisms

    Institute of Scientific and Technical Information of China (English)

    Zhi-yuan Guo; Xun Sun; Xiao-long Xu; Qing Zhao; Jiang Peng; Yu Wang

    2015-01-01

    Human umbilical cord-derived mesenchymal stem cells (hUCMSCs) represent a promising young-state stem cell source for cell-based therapy. hUCMSC transplantation into the transected sciatic nerve promotes axonal regeneration and functional recovery. To further clarify the para-crine effects of hUCMSCs on nerve regeneration, we performed human cytokine antibody array analysis, which revealed that hUCMSCs express 14 important neurotrophic factors. Enzyme-linked immunosorbent assay and immunohistochemistry showed that brain-derived neurotrophic factor, glial-derived neurotrophic factor, hepatocyte growth factor, neurotrophin-3, basic fibroblast growth factor, type I collagen, fibronectin and laminin were highly expressed. Treatment with hUCMSC-conditioned medium enhanced Schwann cell viability and proliferation, increased nerve growth factor and brain-derived neurotrophic factor expression in Schwann cells, and enhanced neurite growth from dorsal root ganglion explants. These ifndings suggest that paracrine action may be a key mechanism underlying the effects of hUCMSCs in peripheral nerve repair.

  3. Human umbilical cord mesenchymal stem cells promote peripheral nerve repair via paracrine mechanisms

    Directory of Open Access Journals (Sweden)

    Zhi-yuan Guo

    2015-01-01

    Full Text Available Human umbilical cord-derived mesenchymal stem cells (hUCMSCs represent a promising young-state stem cell source for cell-based therapy. hUCMSC transplantation into the transected sciatic nerve promotes axonal regeneration and functional recovery. To further clarify the paracrine effects of hUCMSCs on nerve regeneration, we performed human cytokine antibody array analysis, which revealed that hUCMSCs express 14 important neurotrophic factors. Enzyme-linked immunosorbent assay and immunohistochemistry showed that brain-derived neurotrophic factor, glial-derived neurotrophic factor, hepatocyte growth factor, neurotrophin-3, basic fibroblast growth factor, type I collagen, fibronectin and laminin were highly expressed. Treatment with hUCMSC-conditioned medium enhanced Schwann cell viability and proliferation, increased nerve growth factor and brain-derived neurotrophic factor expression in Schwann cells, and enhanced neurite growth from dorsal root ganglion explants. These findings suggest that paracrine action may be a key mechanism underlying the effects of hUCMSCs in peripheral nerve repair.

  4. ENDOCANNABINOIDS INHIBIT RELEASE OF NERVE GROWTH FACTOR BY INFLAMMATION-ACTIVATED MAST CELLS

    OpenAIRE

    2011-01-01

    Abstract Nerve growth factor (NGF) is a pleiotropic member of the neurotrophin family. Beside its neuronal effects, NGF plays a role in various processes, including angiogenesis. Mast cells release NGF and are among elements contributing to angiogenesis, a process regulated by arrays of factors, including the inhibitory cannabinoids. The possible inhibitory role of cannabinoids on mast cell-related NGF mitogenic effect on endothelial cells was then investigated. Human mastocytic ce...

  5. Density-dependent nerve growth factor regulation of Gs-alpha RNA in pheochromocytoma 12 cells.

    Science.gov (United States)

    Tjaden, G; Aguanno, A; Kumar, R; Benincasa, D; Gubits, R M; Yu, H; Dolan, K P

    1990-01-01

    Nerve growth factor (NGF) affects levels of the alpha subunit of the stimulatory G protein (Gs-alpha) in pheochromocytoma 12 cells in a bidirectional, density-dependent manner. Cells grown at high density responded to NGF treatment with increased levels of Gs-alpha mRNA and protein. Conversely, in cells grown in low-density cultures, levels of this mRNA were lowered by NGF treatment. Images PMID:2160599

  6. Amyloid beta resistance in nerve cell lines is mediated by the Warburg effect.

    Directory of Open Access Journals (Sweden)

    Jordan T Newington

    Full Text Available Amyloid beta (Aβ peptide accumulation in the brains of patients with Alzheimer's disease (AD is closely associated with increased nerve cell death. However, many cells survive and it is important to understand the mechanisms involved in this survival response. Recent studies have shown that an anti-apoptotic mechanism in cancer cells is mediated by aerobic glycolysis, also known as the Warburg effect. One of the major regulators of aerobic glycolysis is pyruvate dehydrogenase kinase (PDK, an enzyme which represses mitochondrial respiration and forces the cell to rely heavily on glycolysis, even in the presence of oxygen. Recent neuroimaging studies have shown that the spatial distribution of aerobic glycolysis in the brains of AD patients strongly correlates with Aβ deposition. Interestingly, clonal nerve cell lines selected for resistance to Aβ exhibit increased glycolysis as a result of activation of the transcription factor hypoxia inducible factor 1. Here we show that Aβ resistant nerve cell lines upregulate Warburg effect enzymes in a manner reminiscent of cancer cells. In particular, Aβ resistant nerve cell lines showed elevated PDK1 expression in addition to an increase in lactate dehydrogenase A (LDHA activity and lactate production when compared to control cells. In addition, mitochondrial derived reactive oxygen species (ROS were markedly diminished in resistant but not sensitive cells. Chemically or genetically inhibiting LDHA or PDK1 re-sensitized resistant cells to Aβ toxicity. These findings suggest that the Warburg effect may contribute to apoptotic-resistance mechanisms in the surviving neurons of the AD brain. Loss of the adaptive advantage afforded by aerobic glycolysis may exacerbate the pathophysiological processes associated with AD.

  7. Regulation of Schwann cell proliferation in cultured segments of the adult rat sciatic nerve

    DEFF Research Database (Denmark)

    Svenningsen, Åsa Fex; Kanje, M

    1998-01-01

    Schwann cell proliferation was studied in cultured segments of the rat sciatic nerve by measurement of [3H] thymidine incorporation or through bromodeoxyuridine-(BrdU)-labelling and immunocytochemistry. The aim was to delineate mechanisms involved in the injury-induced proliferative response...

  8. Axotomy induces MHC class I antigen expression on rat nerve cells

    DEFF Research Database (Denmark)

    Maehlen, J; Schröder, H D; Klareskog, L;

    1988-01-01

    Immunomorphological staining demonstrates that class I major histocompatibility complex (MHC)-coded antigen expression can be selectively induced on otherwise class I-negative rat nerve cells by peripheral axotomy. Induction of class I as well as class II antigen expression was simultaneously see...

  9. Bioreactor and methods for producing synchronous cells

    Science.gov (United States)

    Helmstetter, Charles E. (Inventor); Thornton, Maureen (Inventor); Gonda, Steve (Inventor)

    2005-01-01

    Apparatus and methods are directed to a perfusion culture system in which a rotating bioreactor is used to grow cells in a liquid culture medium, while these cells are attached to an adhesive-treated porous surface. As a result of this arrangement and its rotation, the attached cells divide, with one cell remaining attached to the substrate, while the other cell, a newborn cell is released. These newborn cells are of approximately the same age, that are collected upon leaving the bioreactor. The populations of newborn cells collected are of synchronous and are minimally, if at all, disturbed metabolically.

  10. Effect of glial cell line-derived neurotrophic factor on peripheral nerve regeneration in adult rat

    Institute of Scientific and Technical Information of China (English)

    CHEN Zhe-yu; LI Jian-hong; ZHENG Xing-dong; LU Chang-lin; HE Cheng

    2001-01-01

    Objective: To study the effect of glial cell line-derived neurotrophic (GDNF) on adult peripheral nerve regeneration. Methods: Transectioned sciatic nerve in adult rats was sutured into silicone channel. GDNF or SAL solution was injected into the silicone channels during operation. Four weeks later, the effect of GDNF on axonal regeneration was evaluated by degenerative neurofiber staining and HRP retrograde tracing. Results: Compared with SAL group, the percentage of degenerative neurofiber areas decreased from 17.3% to 1.9% ( P<0.01 ) and the ratio of labeled spinal somas number was significantly increased from 43.5% to 68.3% ( P<0.01 ) in GDNF group. Conclusion: The results suggest that exogenous GDNF can obviously enhance adult peripheral nerve regeneration.

  11. Peripheral Neuropathy and VIth Nerve Palsy Related to Randall Disease Successfully Treated by High-Dose Melphalan, Autologous Blood Stem Cell Transplantation, and VIth Nerve Decompression Surgery

    Directory of Open Access Journals (Sweden)

    C. Foguem

    2010-01-01

    Full Text Available Randall disease is an unusual cause of extraocular motor nerve (VI palsy. A 35-year-old woman was hospitalized for sicca syndrome. The physical examination showed general weakness, weight loss, diplopia related to a left VIth nerve palsy, hypertrophy of the submandibular salivary glands, and peripheral neuropathy. The biological screening revealed renal insufficiency, serum monoclonal kappa light chain immunoglobulin, urinary monoclonal kappa light chain immunoglobulin, albuminuria, and Bence-Jones proteinuria. Bone marrow biopsy revealed medullar plasma cell infiltration. Immunofixation associated with electron microscopy analysis of the salivary glands showed deposits of kappa light chains. Randall disease was diagnosed. The patient received high-dose melphalan followed by autostem cell transplantation which led to rapid remission. Indeed, at the 2-month followup assessment, the submandibular salivary gland hypertrophy and renal insufficiency had disappeared, and the peripheral neuropathy, proteinuria, and serum monoclonal light chain had decreased significantly. The persistent diplopia was treated with nerve decompression surgery of the left extraocular motor nerve. Cranial nerve complications of Randall disease deserve to be recognized.

  12. Distribution of mesenchymal stem cells and effects on neuronal survival and axon regeneration after optic nerve crush and cell therapy.

    Directory of Open Access Journals (Sweden)

    Louise Alessandra Mesentier-Louro

    Full Text Available Bone marrow-derived cells have been used in different animal models of neurological diseases. We investigated the therapeutic potential of mesenchymal stem cells (MSC injected into the vitreous body in a model of optic nerve injury. Adult (3-5 months old Lister Hooded rats underwent unilateral optic nerve crush followed by injection of MSC or the vehicle into the vitreous body. Before they were injected, MSC were labeled with a fluorescent dye or with superparamagnetic iron oxide nanoparticles, which allowed us to track the cells in vivo by magnetic resonance imaging. Sixteen and 28 days after injury, the survival of retinal ganglion cells was evaluated by assessing the number of Tuj1- or Brn3a-positive cells in flat-mounted retinas, and optic nerve regeneration was investigated after anterograde labeling of the optic axons with cholera toxin B conjugated to Alexa 488. Transplanted MSC remained in the vitreous body and were found in the eye for several weeks. Cell therapy significantly increased the number of Tuj1- and Brn3a-positive cells in the retina and the number of axons distal to the crush site at 16 and 28 days after optic nerve crush, although the RGC number decreased over time. MSC therapy was associated with an increase in the FGF-2 expression in the retinal ganglion cells layer, suggesting a beneficial outcome mediated by trophic factors. Interleukin-1β expression was also increased by MSC transplantation. In summary, MSC protected RGC and stimulated axon regeneration after optic nerve crush. The long period when the transplanted cells remained in the eye may account for the effect observed. However, further studies are needed to overcome eventually undesirable consequences of MSC transplantation and to potentiate the beneficial ones in order to sustain the neuroprotective effect overtime.

  13. Brain changes in Alzheimer's disease patients with implanted encapsulated cells releasing nerve growth factor.

    Science.gov (United States)

    Ferreira, Daniel; Westman, Eric; Eyjolfsdottir, Helga; Almqvist, Per; Lind, Göran; Linderoth, Bengt; Seiger, Ake; Blennow, Kaj; Karami, Azadeh; Darreh-Shori, Taher; Wiberg, Maria; Simmons, Andrew; Wahlund, Lars-Olof; Wahlberg, Lars; Eriksdotter, Maria

    2015-01-01

    New therapies with disease-modifying effects are urgently needed for treating Alzheimer's disease (AD). Nerve growth factor (NGF) protein has demonstrated regenerative and neuroprotective effects on basal forebrain cholinergic neurons in animal studies. In addition, AD patients treated with NGF have previously shown improved cognition, EEG activity, nicotinic binding, and glucose metabolism. However, no study to date has analyzed brain atrophy in patients treated with NGF producing cells. In this study we present MRI results of the first clinical trial in patients with AD using encapsulated NGF biodelivery to the basal forebrain. Six AD patients received the treatment during twelve months. Patients were grouped as responders and non-responders according to their twelve-months change in MMSE. Normative values were created from 131 AD patients from ADNI, selecting 36 age- and MMSE-matched patients for interpreting the longitudinal changes in MMSE and brain atrophy. Results at baseline indicated that responders showed better clinical status and less pathological levels of cerebrospinal fluid (CSF) Aβ1-42. However, they showed more brain atrophy, and neuronal degeneration as evidenced by higher CSF levels of T-tau and neurofilaments. At follow-up, responders showed less brain shrinkage and better progression in the clinical variables and CSF biomarkers. Noteworthy, two responders showed less brain shrinkage than the normative ADNI group. These results together with previous evidence supports the idea that encapsulated biodelivery of NGF might have the potential to become a new treatment strategy for AD with both symptomatic and disease-modifying effects.

  14. A comparative study of gland cells implicated in the nerve dependence of salamander limb regeneration.

    Science.gov (United States)

    Kumar, Anoop; Nevill, Graham; Brockes, Jeremy P; Forge, Andrew

    2010-07-01

    Limb regeneration in salamanders proceeds by formation of the blastema, a mound of proliferating mesenchymal cells surrounded by a wound epithelium. Regeneration by the blastema depends on the presence of regenerating nerves and in earlier work it was shown that axons upregulate the expression of newt anterior gradient (nAG) protein first in Schwann cells of the nerve sheath and second in dermal glands underlying the wound epidermis. The expression of nAG protein after plasmid electroporation was shown to rescue a denervated newt blastema and allow regeneration to the digit stage. We have examined the dermal glands by scanning and transmission electron microscopy combined with immunogold labelling of the nAG protein. It is expressed in secretory granules of ductless glands, which apparently discharge by a holocrine mechanism. No external ducts were observed in the wound epithelium of the newt and axolotl. The larval skin of the axolotl has dermal glands but these are absent under the wound epithelium. The nerve sheath was stained post-amputation in innervated but not denervated blastemas with an antibody to axolotl anterior gradient protein. This antibody reacted with axolotl Leydig cells in the wound epithelium and normal epidermis. Staining was markedly decreased in the wound epithelium after denervation but not in the epidermis. Therefore, in both newt and axolotl the regenerating axons induce nAG protein in the nerve sheath and subsequently the protein is expressed by gland cells, under (newt) or within (axolotl) the wound epithelium, which discharge by a holocrine mechanism. These findings serve to unify the nerve dependence of limb regeneration.

  15. Hirschsprung's disease: Is there a relationship between mast cells and nerve fibers?

    Institute of Scientific and Technical Information of China (English)

    Amit Kumar Yadav; Kiran Mishra; Anup Mohta; Sarla Agarwal

    2009-01-01

    AIM:To define the topography of mast cells and their numbers in cases of Hirschsprung's disease (HD) and non-HD, assess neural hypertrophy using imaging software and to study the relationship between mast cells and nerve fibers. METHODS:HE stained sections of 32 cases of chronic constipation in the age group of 0-14 years were reviewed for ganglion cells. AChE staining was performed on frozen sections of colonic and rectal biopsies. Based on their findings cases were divided into HD and non-HD and mast cells stained by toluidine blue were evaluated. Image analysis by computerized software was applied to S-100 stained sections for assessment of neural hypertrophy. RESULTS:Difference between number of mast cells in HD group (mean = 36.44) and in non-HD group (mean = 14.79) was statistically significant. Image analysis morphometry on S-100 stained sections served as a useful adjunct. The difference between number, size, and perimeter of the nerve fibers between HD and non-HD group was statistically significant. CONCLUSION:Mast cells are significantly increased in HD and their base line values are much higher in Indian children than that reported in Western literature. Their role in HD needs further research. Morphometry of S-100 stained nerve fibers is a useful adjunct to conventional methods for diagnosis of HD.

  16. Novel Adult Stem Cells for Peripheral Nerve Regeneration

    Science.gov (United States)

    2013-09-01

    derive from hair follicle precursors and exhibit properties of adult dermal stem cells. Cell Stem Cell 5, 610–623 (2009). 53. Morrison, S. J., White...potential therapeutic target of vascular diseases. MVSCs in arteries and veins may have different developmental origins. Wnt1 lineage-tracing experiments...and may lead to new therapies using MVSCs as a therapeutic target . Methods Generation of transgenic mice and genotyping. Animal studies were approved

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

    Directory of Open Access Journals (Sweden)

    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

  18. Engineered neural tissue with aligned, differentiated adipose-derived stem cells promotes peripheral nerve regeneration across a critical sized defect in rat sciatic nerve.

    Science.gov (United States)

    Georgiou, Melanie; Golding, Jon P; Loughlin, Alison J; Kingham, Paul J; Phillips, James B

    2015-01-01

    Adipose-derived stem cells were isolated from rats and differentiated to a Schwann cell-like phenotype in vitro. The differentiated cells (dADSCs) underwent self-alignment in a tethered type-1 collagen gel, followed by stabilisation to generate engineered neural tissue (EngNT-dADSC). The pro-regenerative phenotype of dADSCs was enhanced by this process, and the columns of aligned dADSCs in the aligned collagen matrix supported and guided neurite extension in vitro. EngNT-dADSC sheets were rolled to form peripheral nerve repair constructs that were implanted within NeuraWrap conduits to bridge a 15 mm gap in rat sciatic nerve. After 8 weeks regeneration was assessed using immunofluorescence imaging and transmission electron microscopy and compared to empty conduit and nerve graft controls. The proportion of axons detected in the distal stump was 3.5 fold greater in constructs containing EngNT-dADSC than empty tube controls. Our novel combination of technologies that can organise autologous therapeutic cells within an artificial tissue construct provides a promising new cellular biomaterial for peripheral nerve repair.

  19. Brain-derived neurotrophic factor from bone marrow-derived cells promotes post-injury repair of peripheral nerve.

    Directory of Open Access Journals (Sweden)

    Yoshinori Takemura

    Full Text Available Brain-derived neurotrophic factor (BDNF stimulates peripheral nerve regeneration. However, the origin of BNDF and its precise effect on nerve repair have not been clarified. In this study, we examined the role of BDNF from bone marrow-derived cells (BMDCs in post-injury nerve repair. Control and heterozygote BDNF knockout mice (BDNF+/- received a left sciatic nerve crush using a cerebral blood clip. Especially, for the evaluation of BDNF from BMDCs, studies with bone marrow transplantation (BMT were performed before the injury. We evaluated nerve function using a rotarod test, sciatic function index (SFI, and motor nerve conduction velocity (MNCV simultaneously with histological nerve analyses by immunohistochemistry before and after the nerve injury until 8 weeks. BDNF production was examined by immunohistochemistry and mRNA analyses. After the nerve crush, the controls showed severe nerve dysfunction evaluated at 1 week. However, nerve function was gradually restored and reached normal levels by 8 weeks. By immunohistochemistry, BDNF expression was very faint before injury, but was dramatically increased after injury at 1 week in the distal segment from the crush site. BDNF expression was mainly co-localized with CD45 in BMDCs, which was further confirmed by the appearance of GFP-positive cells in the BMT study. Variant analysis of BDNF mRNA also confirmed this finding. BDNF+/- mice showed a loss of function with delayed histological recovery and BDNF+/+→BDNF+/- BMT mice showed complete recovery both functionally and histologically. These results suggested that the attenuated recovery of the BDNF+/- mice was rescued by the transplantation of BMCs and that BDNF from BMDCs has an essential role in nerve repair.

  20. A study on peripheral nerve regeneration via biomimetic conduits loaded with Schwann cells and nerve growth factor

    Institute of Scientific and Technical Information of China (English)

    ZHAO Fengyi; ZHOU Peilan; WANG Ruilin; YANG Mingfu; ZHAO Weisheng; WEI Dian; ZHANG Tieliang; YAO Kangde; CUI Yuanlu

    2001-01-01

    @@ Guided tissue regeneration is a new approach in the reconstructive surgery of peripheral nerves. Biomimetic conducts were construct from the expanded vein onwhose inner surface composited with amnion filaments (cf. Fig 1).

  1. Characterisation of a Cell Culture System for Investigating Nerve Agent Neurotoxicology. Part 1

    Science.gov (United States)

    2012-03-01

    implications for the mechanisms of toxicity. Biochim. Biophys. Acta 1190, 149-154. Bajgar,J. (2004). Organophosphates /nerve agent poisoning : mechanism of...can produce long-term CNS damage and/or deficits. Although acute OP poisoning is a well- established clinical entity, the existence of chronic... poisoning due to exposure to low levels of OPs (below the threshold required for cholinergic clinical symptoms) is an issue of debate (Pope, 1999

  2. Enhancing nerve regeneration in the peripheral nervous system using polymeric scaffolds, stem cell engineering and nanoparticle delivery system

    Science.gov (United States)

    Sharma, Anup Dutt

    Peripheral nerve regeneration is a complex biological process responsible for regrowth of neural tissue following a nerve injury. The main objective of this project was to enhance peripheral nerve regeneration using interdisciplinary approaches involving polymeric scaffolds, stem cell therapy, drug delivery and high content screening. Biocompatible and biodegradable polymeric materials such as poly (lactic acid) were used for engineering conduits with micropatterns capable of providing mechanical support and orientation to the regenerating axons and polyanhydrides for fabricating nano/microparticles for localized delivery of neurotrophic growth factors and cytokines at the site of injury. Transdifferentiated bone marrow stromal cells or mesenchymal stem cells (MSCs) were used as cellular replacements for lost native Schwann cells (SCs) at the injured nerve tissue. MSCs that have been transdifferentiated into an SC-like phenotype were tested as a substitute for the myelinating SCs. Also, genetically modified MSCs were engineered to hypersecrete brain- derived neurotrophic factor (BDNF) and glial cell line-derived neurotrophic factor (GDNF) to secrete therapeutic factors which Schwann cell secrete. To further enhance the regeneration, nerve growth factor (NGF) and interleukin-4 (IL4) releasing polyanhydrides nano/microparticles were fabricated and characterized in vitro for their efficacy. Synergistic use of these proposed techniques was used for fabricating a multifunctional nerve regeneration conduit which can be used as an efficient tool for enhancing peripheral nerve regeneration.

  3. 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.

  4. 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

    2017-01-01

    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. PMID:28067793

  5. Fetal Nerve Cell Transplantation in Early Post-Resuscitation Period in Rats

    Directory of Open Access Journals (Sweden)

    Damira Tazhibayeva

    2015-02-01

    Full Text Available Introduction. Fetal cell transplantation is a promising biomedical approach for disease treatment; however, the use of fetal cell therapy is still experimental. This research was deemed a necessity to provide evidence-based research for the application of cell transplantation as a treatment method. The aim of this study was to evaluate the effect of fetal nerve cell transplantation in rat survivors (and non-survivors after clinical death by mechanical asphyxia.Methods. 68 white laboratory rats were divided into two groups of identical age and sex: a control group of 12-month adult male rats (n = 26 and an experimental group (n = 42. Rats were fixed under ether anesthesia. We then blocked the oral and nasal regions with cotton wool soaked in saline solution. A four-minute clinical death though acute mechanical asphyxia was simulated by applying the method of N. Shim. After the 4-minute clinical death, we resuscitated the rats using external cardiac massage and artifical respiration. Suspension of the fetal nerve cells was injected intraperitoneally at 1mm3 per 25g at the time of cardiac activity restoration. Lactate dehydrogenase (LDH and creatine phosphokinase (CPK levels were examined in the homogenate cerebral cortex of reanimated animals. We recorded the survival rate during the post-resuscitation period and analyzed the integrative brain functions using anxiety-phobic status and latent inhibition.Results. After fetal nerve cell transplantation, the enzymatic reactions in the experimental group became normal with a significant decrease in LDH and an increase in CPK levels compared to the control group. In the control group, 10 rats died and 16 lived (62% survival rate, while 7 rats died and 35 lived (83% survival rate in the experimental group during the first 7 days. Rats that did not receive the treatment tended to die sooner than those in the experimental group. As a result of transplantation, the anxiety level in the experimental group

  6. Radiosensitivity of glial progenitor cells of the perinatal and adult rat optic nerve studied by an in vitro clonogenic assay

    Energy Technology Data Exchange (ETDEWEB)

    Maazen, R.W.M. van der; Verhagen, I.; Kleiboer, B.J.; Kogel, A.J. van der (Nijmegen University (Netherlands). Institute of Radiotherapy)

    1991-04-01

    The cellular basis of radiation-induced demyelination and white matter necrosis of the central nervous system (CNS), is poorly understood. Glial cells responsible for myelination in the CNS might be the target cells of this type of damage. Glial cells with stem cell properties derived from the perinatal and adult rat CNS can be cultured in vitro. These cells are able to differentiate into oligodendrocytes or type-2 astrocytes (O-2A) depending on the culture conditions. Growth factors produced by monolayers of type-1 astrocytes inhibit premature differentiation of O-2A progenitor cells and allow colony formation. A method which employs these monolayers of type-1 astrocytes to culture O-2A progenitor cells has been adapted to allow the analysis of colonies of surviving cells after X-irradiation. In vitro survival curves were obtained for glial progenitor cells derived from perinatal and adult optic nerves. The intrinsic radiosensitivity of perinatal and adult O-2A progenitor cells showed a large difference. Perinatal O-2A progenitor cells are quite radiosensitive, in contrast to adult O-2A progenitor cells. For both cell types an inverse relationship was found between the dose and the size of colonies derived from surviving cells. Surviving O-2A progenitor cells maintain their ability to differentiate into oligo-dendrocytes or type-2 astrocytes. This system to assess radiation-induced damage to glial progenitor cells in vitro systems to have a great potential in unraveling the cellular basis of radiation-induced demyelinating syndromes of the CNS. (author). 28 refs.; 4 figs.; 1 tab.

  7. Enhancement of Peripheral Nerve Regrowth by the Purine Nucleoside Analog and Cell Cycle Inhibitor, Roscovitine

    Science.gov (United States)

    Law, Vincent; Dong, Sophie; Rosales, Jesusa L.; Jeong, Myung-Yung; Zochodne, Douglas; Lee, Ki-Young

    2016-01-01

    Peripheral nerve regeneration is a slow process that can be associated with limited outcomes and thus a search for novel and effective therapy for peripheral nerve injury and disease is crucial. Here, we found that roscovitine, a synthetic purine nucleoside analog, enhances neurite outgrowth in neuronal-like PC12 cells. Furthermore, ex vivo analysis of pre-injured adult rat dorsal root ganglion (DRG) neurons showed that roscovitine enhances neurite regrowth in these cells. Likewise, in vivo transected sciatic nerves in rats locally perfused with roscovitine had augmented repopulation of new myelinated axons beyond the transection zone. By mass spectrometry, we found that roscovitine interacts with tubulin and actin. It interacts directly with tubulin and causes a dose-dependent induction of tubulin polymerization as well as enhances Guanosine-5′-triphosphate (GTP)-dependent tubulin polymerization. Conversely, roscovitine interacts indirectly with actin and counteracts the inhibitory effect of cyclin-dependent kinases 5 (Cdk5) on Actin-Related Proteins 2/3 (Arp2/3)-dependent actin polymerization, and thus, causes actin polymerization. Moreover, in the presence of neurotrophic factors such as nerve growth factor (NGF), roscovitine-enhanced neurite outgrowth is mediated by increased activation of the extracellular signal-regulated kinases 1/2 (ERK1/2) and p38 mitogen-activated protein kinase (MAPK) pathways. Since microtubule and F-actin dynamics are critical for axonal regrowth, the ability of roscovitine to activate the ERK1/2 and p38 MAPK pathways and support polymerization of tubulin and actin indicate a major role for this purine nucleoside analog in the promotion of axonal regeneration. Together, our findings demonstrate a therapeutic potential for the purine nucleoside analog, roscovitine, in peripheral nerve injury.

  8. JNK3 involvement in nerve cell apoptosis and neurofunctional recovery after traumatic brain injury☆

    Institute of Scientific and Technical Information of China (English)

    Jiang Long; Li Cai; Jintao Li; Lei Zhang; Haiyang Yang; Tinghua Wang

    2013-01-01

    Increasing evidence has revealed that the activation of the JNK pathway participates in apoptosis of nerve cells and neurological function recovery after traumatic brain injury. However, which genes in the JNK family are activated and their role in traumatic brain injury remain unclear. Therefore, in this study, in situ end labeling, reverse transcription-PCR and neurological function assessment were adopted to investigate the alteration of JNK1, JNK2 and JNK3 gene expression in cerebral injured rats, and their role in cell apoptosis and neurological function restoration. Results showed that JNK3 expression significantly decreased at 1 and 6 hours and 1 and 7 days post injury, but that JNK1 and JNK2 expression remained unchanged. In addition, the number of apoptotic nerve cells surrounding the injured cerebral cortex gradually reduced over time post injury. The Neurological Severity Scores gradually decreased over 1, 3, 5, 14 and 28 days post injury. These findings suggested that JNK3 expression was downregulated at early stages of brain injury, which may be associated with apoptosis of nerve cells. Downregulation of JNK3 expression may promote the recovery of neurological function following traumatic brain injury.

  9. Current progress in use of adipose derived stem cells inperipheral nerve regeneration

    Institute of Scientific and Technical Information of China (English)

    Shomari DL Zack-Williams; Peter E Butler; Deepak M Kalaskar

    2015-01-01

    Unlike central nervous system neurons; those in theperipheral nervous system have the potential for fullregeneration after injury. Following injury, recovery iscontrolled by schwann cells which replicate and modulatethe subsequent immune response. The level of nerverecovery is strongly linked to the severity of the initialinjury despite the significant advancements in imagingand surgical techniques. Multiple experimental modelshave been used with varying successes to augment thenatural regenerative processes which occur following nerveinjury. Stem cell therapy in peripheral nerve injury maybe an important future intervention to improve the bestattainable clinical results. In particular adipose derivedstem cells (ADSCs) are multipotent mesenchymal stemcells similar to bone marrow derived stem cells, which arethought to have neurotrophic properties and the ability todifferentiate into multiple lineages. They are ubiquitouswithin adipose tissue; they can form many structuresresembling the mature adult peripheral nervous system.Following early in vitro work; multiple small and largeanimal in vivo models have been used in conjunction withconduits, autografts and allografts to successfully bridgethe peripheral nerve gap. Some of the ADSC relatedneuroprotective and regenerative properties have beenelucidated however much work remains before a modelcan be used successfully in human peripheral nerve injury(PNI). This review aims to provide a detailed overview ofprogress made in the use of ADSC in PNI, with discussionon the role of a tissue engineered approach for PNI repair.

  10. Biological conduits combining bone marrow mesenchymal stem cells and extracellular matrix to treat long-segment sciatic nerve defects

    Institute of Scientific and Technical Information of China (English)

    Yang Wang; Zheng-wei Li; Min Luo; Ya-jun Li; Ke-qiang Zhang

    2015-01-01

    The transplantation of polylactic glycolic acid conduits combining bone marrow mesenchymal stem cells and extracellular matrix gel for the repair of sciatic nerve injury is effective in some re-spects, but few data comparing the biomechanical factors related to the sciatic nerve are available. In the present study, rabbit models of 10-mm sciatic nerve defects were prepared. The rabbit models were repaired with autologous nerve, a polylactic glycolic acid conduit+bone marrow mesenchymal stem cells, or a polylactic glycolic acid conduit+bone marrow mesenchymal stem cells+extracellular matrix gel. After 24 weeks, mechanical testing was performed to determine the stress relaxation and creep parameters. Following sciatic nerve injury, the magnitudes of the stress decrease and strain increase at 7,200 seconds were largest in the polylactic glycolic acid conduit+bone marrow mesenchymal stem cells+extracellular matrix gel group, followed by the polylactic glycolic acid conduit+bone marrow mesenchymal stem cells group, and then the autologous nerve group. Hematoxylin-eosin staining demonstrated that compared with the poly-lactic glycolic acid conduit+bone marrow mesenchymal stem cells group and the autologous nerve group, a more complete sciatic nerve regeneration was found, including good myelination, regularly arranged nerve ifbers, and a completely degraded and resorbed conduit, in the polylac-tic glycolic acid conduit+bone marrow mesenchymal stem cells+extracellular matrix gel group. These results indicate that bridging 10-mm sciatic nerve defects with a polylactic glycolic acid conduit+bone marrow mesenchymal stem cells+extracellular matrix gel construct increases the stress relaxation under a constant strain, reducing anastomotic tension. Large elongations under a constant physiological load can limit the anastomotic opening and shift, which is ben-eifcial for the regeneration and functional reconstruction of sciatic nerve. Better regeneration was found with the

  11. Effect of Human Cytomegalovirus Infection on Nerve Growth Factor Expression in Human Glioma U251 Cells

    Institute of Scientific and Technical Information of China (English)

    HAI-TAO WANG; BIN WANG; ZHI-JUN LIU; ZHI-QIANG BAI; LING LI; HAI-YAN LIU; DONG-MENG QIAN; ZHI-YONG YAN; XU-XIA SONG

    2009-01-01

    Objectives To explore the change of endogenic nerve growth factor (NGF) expression in human glioma cells infected with human cytomegalovirus (HCMV). Methods U251 cells were cultured in RPMI 1640 culture medium and infected with HCMV AD169 strain in vitro to establish a cell model of viral infection. Morphologic changes of U251 cells were observed under inverted microscope before and after infection with HCMV. Expression of NGF gene and protein of cells was detected by RT-PCR and Western blotting before and after infection with HCMV. Results The cytopathic effects of HCMV-infected cells appeared on day 5 after infection. However, differential NGF expression was evident on day 7. NGF expression was decreased significantly in U251 cells on day 7 after infection in comparison with control group (P<0.05). Conclusion HCMV can down-regulate endogenous NGF levels in human glioma cell line U251.

  12. Influence of ketamine on amino acid neurotransmitters secretion by nerve cells in vitro

    Directory of Open Access Journals (Sweden)

    Mingxian Shi

    2016-04-01

    Full Text Available In order to study the influence of amino acid neurotransmitters secreted by the nerve cells after ketamine treatment, the nerve cells were cultured in vitro to exclude the interference of other factors in vivo and treated with three different doses of ketamine (1, 3 and 5 µg/mL. Then, the concentration of neuronal amino acid neurotransmitters was examined at 0, 15, 30, 45, 60, 90, 120 min after treatment. The trends of each amino acid concentration after ketamine treatment were nearly the same among the different treatment doses. After 15 min of adapting time, ketamine decreased the excitatory amino acid glutamic acid and aspartic acid concentration, and increased the concentration of the inhibitory amino acid glycine. Their concentrations showed a tendency to return approximately to the original level after 120 min.

  13. Poly(amidoamine Hydrogels as Scaffolds for Cell Culturing and Conduits for Peripheral Nerve Regeneration

    Directory of Open Access Journals (Sweden)

    Fabio Fenili

    2011-01-01

    Full Text Available Biodegradable and biocompatible poly(amidoamine-(PAA- based hydrogels have been considered for different tissue engineering applications. First-generation AGMA1 hydrogels, amphoteric but prevailing cationic hydrogels containing carboxylic and guanidine groups as side substituents, show satisfactory results in terms of adhesion and proliferation properties towards different cell lines. Unfortunately, these hydrogels are very swellable materials, breakable on handling, and have been found inadequate for other applications. To overcome this problem, second-generation AGMA1 hydrogels have been prepared adopting a new synthetic method. These new hydrogels exhibit good biological properties in vitro with satisfactory mechanical characteristics. They are obtained in different forms and shapes and successfully tested in vivo for the regeneration of peripheral nerves. This paper reports on our recent efforts in the use of first-and second-generation PAA hydrogels as substrates for cell culturing and tubular scaffold for peripheral nerve regeneration.

  14. Composition of Mineral Produced by Dental Mesenchymal Stem Cells.

    Science.gov (United States)

    Volponi, A A; Gentleman, E; Fatscher, R; Pang, Y W Y; Gentleman, M M; Sharpe, P T

    2015-11-01

    Mesenchymal stem cells isolated from different dental tissues have been described to have osteogenic/odontogenic-like differentiation capacity, but little attention has been paid to the biochemical composition of the material that each produces. Here, we used Raman spectroscopy to analyze the mineralized materials produced in vitro by different dental cell populations, and we compared them with the biochemical composition of native dental tissues. We show that different dental stem cell populations produce materials that differ in their mineral and matrix composition and that these differ from those of native dental tissues. In vitro, BCMP (bone chip mass population), SCAP (stem cells from apical papilla), and SHED (stem cells from human-exfoliated deciduous teeth) cells produce a more highly mineralized matrix when compared with that produced by PDL (periodontal ligament), DPA (dental pulp adult), and GF (gingival fibroblast) cells. Principal component analyses of Raman spectra further demonstrated that the crystallinity and carbonate substitution environments in the material produced by each cell type varied, with DPA cells, for example, producing a more carbonate-substituted mineral and with SCAP, SHED, and GF cells creating a less crystalline material when compared with other dental stem cells and native tissues. These variations in mineral composition reveal intrinsic differences in the various cell populations, which may in turn affect their specific clinical applications.

  15. A Novel Model for Acute Peripheral Nerve Injury in the Horse and Evaluation of the Effect of Mesenchymal Stromal Cells Applied In Situ on Nerve Regeneration: A Preliminary Study

    Science.gov (United States)

    Cruz Villagrán, Claudia; Schumacher, Jim; Donnell, Robert; Dhar, Madhu S.

    2016-01-01

    Transplantation of mesenchymal stromal cells (MSCs) to sites of experimentally created nerve injury in laboratory animals has shown promising results in restoring nerve function. This approach for nerve regeneration has not been reported in horses. In this study, we first evaluated the in vitro ability of equine bone marrow-derived MSCs (EBM-MSCs) to trans-differentiate into Schwann-like cells and subsequently tested the MSCs in vivo for their potential to regenerate a transected nerve after implantation. The EBM-MSCs from three equine donors were differentiated into SCLs for 7 days, in vitro, in the presence of specialized differentiation medium and evaluated for morphological characteristics, by using confocal microscopy, and for protein characteristics, by using selected Schwann cell markers (GFAP and S100b). The EBM-MSCs were then implanted into the fascia surrounding the ramus communicans of one fore limb of three healthy horses after a portion of this nerve was excised. The excised portion of the nerve was examined histologically at the time of transection, and stumps of the nerve were examined histologically at day 45 after transplantation. The EBM-MSCs from all donors demonstrated morphological and protein characteristics of those of Schwann cells 7 days after differentiation. Nerves implanted with EBM-MSCs after nerve transection did not show evidence of nerve regeneration at day 45. Examination of peripheral nerves collected 45 days after injury and stem cell treatment revealed no histological differences between nerves treated with MSCs and those treated with isotonic saline solution (controls). The optimal delivery of MSCs and the model suitable to study the efficacy of MSCs in nerve regeneration should be investigated. PMID:27695697

  16. Wide distribution of immunoreactive renin in nerve cells of human brain.

    OpenAIRE

    1980-01-01

    By use of the indirect peroxidase-antiperoxidase complex immunocytochemical technique, antibody to purified human renal renin was applied to formalin-fixed paraffin sections of human cadaver brain. Immune reaction products were observed in most nerve cells in all areas of the brain examined; staining was limited to the soma and proximal dendrites. These experiments have confirmed the presence of a renin-like substance in central nervous tissue and suggest a more generalized function for "brai...

  17. Cells that emerge from embryonic explants produce fibers of type IV collagen.

    Science.gov (United States)

    Chen, J M; Little, C D

    1985-10-01

    Double immunofluorescence staining experiments designed to examine the synthesis and deposition of collagen types I and IV in cultured explants of embryonic mouse lung revealed the presence of connective tissue-like fibers that were immunoreactive with anti-type IV collagen antibodies. This observation is contrary to the widely accepted belief that type IV collagen is found only in sheet-like arrangements beneath epithelia or as a sheath-like layer enveloping bundles of nerve or muscle cells. The extracellular matrix produced by cells that migrate from embryonic mouse lung rudiments in vitro was examined by double indirect immunofluorescence microscopy. Affinity-purified monospecific polyclonal antibodies were used to examine cells after growth on glass or native collagen substrata. The data show that embryonic mesenchymal cells can produce organized fibers of type IV collagen that are not contained within a basement membrane, and that embryonic epithelial cells deposit fibers and strands of type IV collagen beneath their basal surface when grown on glass; however, when grown on a rat tail collagen substratum the epithelial cells produce a fine meshwork. To our knowledge this work represents the first report that type IV collagen can be organized by cells into a fibrous extracellular matrix that is not a basement membrane.

  18. Mesenchymal stem cell therapy in retinal and optic nerve diseases: An update of clinical trials

    Science.gov (United States)

    Labrador-Velandia, Sonia; Alonso-Alonso, María Luz; Alvarez-Sanchez, Sara; González-Zamora, Jorge; Carretero-Barrio, Irene; Pastor, José Carlos; Fernandez-Bueno, Iván; Srivastava, Girish Kumar

    2016-01-01

    Retinal and optic nerve diseases are degenerative ocular pathologies which lead to irreversible visual loss. Since the advanced therapies availability, cell-based therapies offer a new all-encompassing approach. Advances in the knowledge of neuroprotection, immunomodulation and regenerative properties of mesenchymal stem cells (MSCs) have been obtained by several preclinical studies of various neurodegenerative diseases. It has provided the opportunity to perform the translation of this knowledge to prospective treatment approaches for clinical practice. Since 2008, several first steps projecting new treatment approaches, have been taken regarding the use of cell therapy in patients with neurodegenerative pathologies of optic nerve and retina. Most of the clinical trials using MSCs are in I/II phase, recruiting patients or ongoing, and they have as main objective the safety assessment of MSCs using various routes of administration. However, it is important to recognize that, there is still a long way to go to reach clinical trials phase III-IV. Hence, it is necessary to continue preclinical and clinical studies to improve this new therapeutic tool. This paper reviews the latest progress of MSCs in human clinical trials for retinal and optic nerve diseases. PMID:27928464

  19. Substance P combined with epidermal stem cells promotes wound healing and nerve regeneration in diabetes mellitus

    Institute of Scientific and Technical Information of China (English)

    Fei-bin Zhu; Xiang-jing Fang; De-wu Liu; Ying Shao; Hong-yan Zhang; Yan Peng; Qing-ling Zhong; Yong-tie Li; De-ming Liu

    2016-01-01

    Exogenous substance P accelerates wound healing in diabetes, but the mechanism remains poorly understood. Here, we established a rat model by intraperitoneally injecting streptozotocin. Four wounds (1.8 cm diameter) were drilled using a self-made punch onto the back, bilateral to the vertebral column, and then treated using amniotic membrane with epidermal stem cells and/or substance P around and in the middle of the wounds. With the combined treatment the wound-healing rate was 100% at 14 days. With prolonged time, type I col-lagen content gradually increased, yet type III collagen content gradually diminished. Abundant protein gene product 9.5- and substance P-immunoreactive nerve ifbers regenerated. Partial nerve ifber endings extended to the epidermis. The therapeutic effects of combined substance P and epidermal stem cells were better than with amniotic membrane and either factor alone. Our results suggest that the com-bination of substance P and epidermal stem cells effectively contributes to nerve regeneration and wound healing in diabetic rats.

  20. Survival and migration of Schwann cells after the vascularized peripheral nerve grafted into spinal cord in rats

    Institute of Scientific and Technical Information of China (English)

    GUO Qing-shan; WANG Ai-min; WANG Xiao-jun; SUN Hong-zhen; DU Quan-yin

    2005-01-01

    Objective:To study the survival and ability of inducing axonal regeneration of the Schwann cells after the peripheral nerve being grafted into spinal cord. Methods:A total of 30 adult female Wistar rats were randomly divided into the VN (vascularized peripheral nerve) and PN (peripheral nerve) groups. A 5-mm spinal cord defect of the left posterior column was made at the T1-3 vertebral level. The defect was grafted with the vascularized or isolated peripheral nerve respectively. The survival and proliferation of the Schwann cells were assessed by histological and morphometric analysis 8 weeks after the operation. Results:In the VN group, the peripheral nerve grew into the cord with lots of Schwann cells survived and proliferated, and had more NF and S-100 positive fibers than in the PN group. Conclusion:The vascularized peripheral nerve enhances the survival and proliferation of the Schwann cells and prompts the regeneration of injured axon of the central nerve system to certain degree.

  1. Enhancement of Median Nerve Regeneration by Mesenchymal Stem Cells Engraftment in an Absorbable Conduit: Improvement of Peripheral Nerve Morphology with Enlargement of Somatosensory Cortical Representation.

    Directory of Open Access Journals (Sweden)

    Julia Teixeira Oliveira

    2014-10-01

    Full Text Available We studied the morphology and the cortical representation of the median nerve (MN, 10 weeks after a transection immediately followed by treatment with tubulization using a polycaprolactone (PCL conduit with or without bone marrow-derived mesenchymal stem cell (MSC transplant. In order to characterize the cutaneous representation of MN inputs in primary somatosensory cortex (S1, electrophysiological cortical mapping of the somatosensory representation of the forepaw and adjacent body parts was performed after acute lesion of all brachial plexus nerves, except for the MN. This was performed in ten adult male Wistar rats randomly assigned in 3 groups: MN Intact (n=4, PCL-Only (n=3 and PCL+MSC (n=3. Ten weeks before mapping procedures in animals from PCL-Only and PCL+MSC groups, animal were subjected to MN transection with removal of a 4-mm-long segment, immediately followed by suturing a PCL conduit to the nerve stumps with (PCL+MSC group or without (PCL-Only group injection of MSC into the conduit. After mapping the representation of the MN in S1, animals had a segment of the regenerated nerve processed for light and transmission electron microscopy. For histomorphometric analysis of the nerve segment, sample size was increased to 5 animals per experimental group. The PCL+MSC group presented a higher number of myelinated fibers and a larger cortical representation of MN inputs in S1 (3,383±390 fibers; 2.3 mm2, respectively than the PCL-Only group (2,226±575 fibers; 1.6 mm2. In conclusion, MSC-based therapy associated with PCL conduits can improve MN regeneration. This treatment seems to rescue the nerve representation in S1, thus minimizing the stabilization of new representations of adjacent body parts in regions previously responsive to the MN.

  2. Co-transplantation of macaque autologous Schwann cells and human embryonic nerve stem cells in treatment of macaque Parkinson's disease

    Institute of Scientific and Technical Information of China (English)

    Ying Xia; Chengchuan Jiang; Zuowei Cao; Keshan Shi; Yang Wang

    2012-01-01

    Objective:To investigate the therapeutic effects of co-transplantation with Schwann cells (SCs) and human embryonic nerve stem cells (NSCs) on macaque Parkinson's disease (PD). Methods:Macaque autologous SCs and human embryonic NSCs were adopted for the treatment of macaque PD. Results: Six months after transplantation, positron emission computerized tomography showed that 18F-FP-β-CIT was significantly concentrated in the injured striatum in the co-transplanted group. Immunohistochemical staining of transplanted area tissue showed migration of tyroxine hydroxylase positive cells from the transplant area to the surrounding area was significantly increased in the co-transplanted group. Conclusions: Co-transplantation of SCs and NSCs could effectively cure PD in macaques. SCs harvested from the autologous peripheral nerves can avoid rejection and the ethics problems, so it is expected to be applied clinically.

  3. Human umbilical cord mesenchymal stem cell-loaded amniotic membrane for the repair of radial nerve injury

    Institute of Scientific and Technical Information of China (English)

    Zhi Li; Hanjiao Qin; Zishan Feng; Wei Liu; Ye Zhou; Lifeng Yang; Wei Zhao; Youjun Li

    2013-01-01

    In this study, we loaded human umbilical cord mesenchymal stem cells onto human amniotic membrane with epithelial cells to prepare nerve conduits, i.e., a relatively closed nerve regeneration chamber. After neurolysis, the injured radial nerve was enwrapped with the prepared nerve conduit, which was fixed to the epineurium by sutures, with the cellon the inner surface of the conduit. Simultaneously, a 1.0 mL aliquot of human umbilical cord mesenchymal stem cellsuspension was injected into the distal and proximal ends of the injured radial nerve with 1.0 cm intervals. A total of 1.75 × 107 cells were seeded on the amniotic membrane. In the control group, patients received only neurolysis. At 12 weeks after celltransplantation, more than 80%of patients exhibited obvious improvements in muscular strength, and touch and pain sensations. In contrast, these improve-ments were observed only in 55-65% of control patients. At 8 and 12 weeks, muscular electro-physiological function in the region dominated by the injured radial nerve was significantly better in the transplantation group than the control group. After celltransplantation, no immunological rejec-tions were observed. These findings suggest that human umbilical cord mesenchymal stem cel-loaded amniotic membrane can be used for the repair of radial nerve injury.

  4. Bone Marrow-Derived Cells as a Therapeutic Approach to Optic Nerve Diseases

    Directory of Open Access Journals (Sweden)

    Louise A. Mesentier-Louro

    2016-01-01

    Full Text Available Following optic nerve injury associated with acute or progressive diseases, retinal ganglion cells (RGCs of adult mammals degenerate and undergo apoptosis. These diseases have limited therapeutic options, due to the low inherent capacity of RGCs to regenerate and due to the inhibitory milieu of the central nervous system. Among the numerous treatment approaches investigated to stimulate neuronal survival and axonal extension, cell transplantation emerges as a promising option. This review focuses on cell therapies with bone marrow mononuclear cells and bone marrow-derived mesenchymal stem cells, which have shown positive therapeutic effects in animal models of optic neuropathies. Different aspects of available preclinical studies are analyzed, including cell distribution, potential doses, routes of administration, and mechanisms of action. Finally, published and ongoing clinical trials are summarized.

  5. Bone Marrow-Derived Cells as a Therapeutic Approach to Optic Nerve Diseases

    Science.gov (United States)

    Mesentier-Louro, Louise A.; Zaverucha-do-Valle, Camila; Rosado-de-Castro, Paulo H.; Silva-Junior, Almir J.; Pimentel-Coelho, Pedro M.; Mendez-Otero, Rosalia; Santiago, Marcelo F.

    2016-01-01

    Following optic nerve injury associated with acute or progressive diseases, retinal ganglion cells (RGCs) of adult mammals degenerate and undergo apoptosis. These diseases have limited therapeutic options, due to the low inherent capacity of RGCs to regenerate and due to the inhibitory milieu of the central nervous system. Among the numerous treatment approaches investigated to stimulate neuronal survival and axonal extension, cell transplantation emerges as a promising option. This review focuses on cell therapies with bone marrow mononuclear cells and bone marrow-derived mesenchymal stem cells, which have shown positive therapeutic effects in animal models of optic neuropathies. Different aspects of available preclinical studies are analyzed, including cell distribution, potential doses, routes of administration, and mechanisms of action. Finally, published and ongoing clinical trials are summarized. PMID:26649049

  6. Bridging the gap between postembryonic cell lineages and identified embryonic neuroblasts in the ventral nerve cord of Drosophila melanogaster

    Directory of Open Access Journals (Sweden)

    Oliver Birkholz

    2015-03-01

    Full Text Available The clarification of complete cell lineages, which are produced by specific stem cells, is fundamental for understanding mechanisms, controlling the generation of cell diversity and patterning in an emerging tissue. In the developing Central Nervous System (CNS of Drosophila, neural stem cells (neuroblasts exhibit two periods of proliferation: During embryogenesis they produce primary lineages, which form the larval CNS. After a phase of mitotic quiescence, a subpopulation of them resumes proliferation in the larva to give rise to secondary lineages that build up the CNS of the adult fly. Within the ventral nerve cord (VNC detailed descriptions exist for both primary and secondary lineages. However, while primary lineages have been linked to identified neuroblasts, the assignment of secondary lineages has so far been hampered by technical limitations. Therefore, primary and secondary neural lineages co-existed as isolated model systems. Here we provide the missing link between the two systems for all lineages in the thoracic and abdominal neuromeres. Using the Flybow technique, embryonic neuroblasts were identified by their characteristic and unique lineages in the living embryo and their further development was traced into the late larval stage. This comprehensive analysis provides the first complete view of which embryonic neuroblasts are postembryonically reactivated along the anterior/posterior-axis of the VNC, and reveals the relationship between projection patterns of primary and secondary sublineages.

  7. Effects of nerve growth factor on the expression of caspase-12 of nerve cells in cerebral ischemia/reperfusion area

    Institute of Scientific and Technical Information of China (English)

    Jiping Yang; Huaijun Liu; Ying Li; Yan Liu; Haiqing Yang

    2006-01-01

    absorbency (A value)was compared with image analytic system; apoptosis rate and apoptosis quantity of nerves in ischemia/reperfusion area were detected with flow cytometry and DNA TdT-mediated biotinylated-Dutp nick end lawith q test every two groups.MATN OUTCOME MEASURES: Expressions of caspase-12 and caspase-3 and apoptosis in cerebral ischemia/reperfusion area in the 3 groups.RESULTS: All 26 rabbits were involved in the final analysis. ① Expressions of caspase-12 and caspase-3:Expression of caspase-12 was 0.36±0.02 in I/R group and 0.13±0.03 in treatment group; expression of caspase-3 was 0.49±0.05 and 0.27±0.06, respectively. Both of them were higher than those in sham operation group (0.07±0.02, 0.09±0.03, P < 0.01), and expressions of two proteases were lower in treatment group than those in I/R group (P < 0.01). There were significant differences of expression of caspase between I/R group and treatment group as compared with that in sham operation group; meanwhile, there (20.2±1.3)% in I/R group and (7.7±0.8)% in treatment group; apoptosis quantity was (32.8±2.6), (7.6±1.5)/high sight, respectively. Both of them were higher and more than those in sham operation group [(4.8±0.4)%, (0.7±0.2) /high sight, P < 0.01]. Apoptosis rate and apoptosis quantity were lower and less in treatment group than those in I/R group (P < 0.01). There was significant difference between I/R group and treatment group as compared with sham operation group, and there was significant difference between treatment group and I/R group.CONCLUSTON: NGF can decrease the number of apoptotic cells of the cerebral ischemia/reperfusion and inhibit the caspase cascade reaction induced by caspase-12, which is one of the protective mechanisms of NGF.

  8. GD3+ cells in the adult rat optic nerve are ramified microglia rather than O-2Aadult progenitor cells.

    Science.gov (United States)

    Wolswijk, G

    1994-04-01

    The adult central nervous system (CNS) contains a population of adult oligodendrocyte-type-2 astrocyte (O-2A) progenitor cells (O-2Aadult progenitor cells). These cells may provide a source of the new oligodendrocytes that are needed to repair demyelinated lesions. In order to examine the role of O-2Aadult progenitor cells in the regeneration of the oligodendrocyte population following demyelinating damage, it is essential to be able to identify such cells unambiguously in sections of adult CNS tissue. The present study examined whether antibodies to the ganglioside GD3 specifically label O-2Aadult progenitor cells in cultures and sections of adult optic nerve, since previous studies on the developing CNS had suggested that O-2Aperinatal progenitor cells were GD3+ in vitro and in vivo. Evidence is presented indicating that, although O-2Aadult progenitor cells in vitro were labelled with the R24 mAb (an anti-GD3 mAb), all GD3+ cells in sections of adult optic nerve bound the OX-42 mAb and the B4 isolectin derived from Griffonia Simplicifolia, and thus were not O-2Aadult progenitor cells, but ramified microglia. The data suggest that O-2Aadult progenitor cells become GD3+ when placed in culture and that ramified microglia lose GD3-expression in vitro.

  9. Salidroside promotes peripheral nerve regeneration based on tissue engineering strategy using Schwann cells and PLGA: in vitro and in vivo

    Science.gov (United States)

    Liu, Hui; Lv, Peizhen; Zhu, Yongjia; Wu, Huayu; Zhang, Kun; Xu, Fuben; Zheng, Li; Zhao, Jinmin

    2017-01-01

    Salidriside (SDS), a phenylpropanoid glycoside derived from Rhodiola rosea L, has been shown to be neuroprotective in many studies, which may be promising in nerve recovery. In this study, the neuroprotective effects of SDS on engineered nerve constructed by Schwann cells (SCs) and Poly (lactic-co-glycolic acid) (PLGA) were studied in vitro. We further investigated the effect of combinational therapy of SDS and PLGA/SCs based tissue engineering on peripheral nerve regeneration based on the rat model of nerve injury by sciatic transection. The results showed that SDS dramatically enhanced the proliferation and function of SCs. The underlying mechanism may be that SDS affects SCs growth through the modulation of neurotrophic factors (BDNF, GDNF and CNTF). 12 weeks after implantation with a 12 mm gap of sciatic nerve injury, SDS-PLGA/SCs achieved satisfying outcomes of nerve regeneration, as evidenced by morphological and functional improvements upon therapy by SDS, PLGA/SCs or direct suture group assessed by sciatic function index, nerve conduction assay, HE staining and immunohistochemical analysis. Our results demonstrated the significant role of introducing SDS into neural tissue engineering to promote nerve regeneration.

  10. Up-regulation of P2X7 receptors mediating proliferation of Schwann cells after sciatic nerve injury.

    Science.gov (United States)

    Song, Xian-min; Xu, Xiao-hui; Zhu, Jiao; Guo, Zhili; Li, Jian; He, Cheng; Burnstock, Geoffrey; Yuan, Hongbin; Xiang, Zhenghua

    2015-06-01

    Peripheral nerve injury (PNI) is a common disease, which results in a partial or total loss of motor, sensory and autonomic functions, leading to a decrease in quality of life. Schwann cells play a vital role in maintaining the peripheral nervous system and in injury and repair. Using immunohistochemistry, Western blot, calcium assay and bromodeoxyuridine (BrdU) proliferation assay, the present study clearly demonstrated that P2X7 receptors (R) were expressed in myelinating and non-myelinating Schwann cells in longitudinal sections of sciatic nerves. After sciatic nerve injury (SNI), P2X7R expression in Schwann cells of injured sciatic nerves was significantly up-regulated during the early days of SNI. Double immunofluorescence of proliferating cell nuclear antigen (PCNA) and P2X7R implied that P2X7R may be involved in proliferation of Schwann cells. Further experiments on primary cultures of Schwann cells showed that P2X7R are functionally expressed in Schwann cells of rat sciatic nerves; ATP via P2X7R can promote Schwann cell proliferation, possibly via the MAPK/ERK intracellular signalling pathway. Other possible roles of P2X7R on Schwann cells are discussed.

  11. Protection of Acanthopanax Senticosus Saponin on Free Radical Injury Induced Aging of Nerve Cell

    Institute of Scientific and Technical Information of China (English)

    潘永进; 顾永健; 顾小苏

    2002-01-01

    Objective: To study the effect of Acanthopanax senticosus saponin (ASS) on free radical injury induced neuron aging. Methods: On day 7 of fetal mice, cortical neuron primary passage cultures were divided into the normal control group, model group and ASS groups. The model group using free radical (FeSO4 plus H2O2) injury mode prepared in vivo cultured ICR mice cortical neuron aging model; ASS groups: 24 hrs before and after treated with H2O2 and FeSO4, different concentration of ASS was added, according to biochemical parameters such as lactate dehydrogenase (LDH), superoxide dismutase (SOD) and malondialdehyde (MDA) etc. and histomorphologic change to observe the protection of ASS on aging neurons. Results: The LDH, SOD, MDA of the model group were compared with the normal group, P<0.01; ASS groups added 1.25 mg/100 ml, 2.5 mg/100 ml, 5 mg/100 ml concentration of ASS, their LDH, SOD, MDA compared with the model group P<0.05-0.01, the difference was significant. In medicated groups the SOD activity of oxidization injured nerve cells obviously elevated, LDH activity and MDA content apparently lowered. Microscope and scanning electron microscopic observation showed that supplemented with ASS to protect the nerve cell injury abated, part of the cellular structure tended to normalize. Conclusion: ASS could act against free radical toxic effect, increase the anti-oxidase activity, strengthen the protection of neuron cells. It is assumed that the effect against nerve cell aging was possibly through scavenging oxygen free radical, strengthening the stability of cell membrane, thus delaying the development of aging.

  12. IL-10-Producing Type 1 Regulatory T Cells and Allergy

    Institute of Scientific and Technical Information of China (English)

    Kui Wu; Yutian Bi; Kun Sun; Changzheng Wang

    2007-01-01

    As an important subset of regulatory T (Treg) cells, IL-10-producing type 1 regulatory T cells (Tr1), have some different features to thymic-derived naturally occurring CD4+CD25+Foxp3+ Treg cells(nTreg cells). Similar to nTreg cells, Tr1 also play important roles in the control of allergic inflammation in several ways. There is a fine balance between Tr1 and Th2 responses in healthy subjects. Skewing of allergic-specific effctor T cells to a Tr1 phenotype appears to be a critical event in successful allergen-specific immunotherapy and glucocorticoids and β2-agonists treatment. Tr1 suppress Th2 cells and effector cells of allergic inflammation, such as eosinophils, mast cells, basophils, through producing IL-10, and perhaps TGF-β. Understanding of Tr1 may be helpful in developing new strategies for treatment of allergic diseases.

  13. Human Adenomyosis Endometrium Stromal Cells Secreting More Nerve Growth Factor: Impact and Effect.

    Science.gov (United States)

    Li, Yan; Zou, Shien; Xia, Xian; Zhang, Shaofen

    2015-09-01

    Abnormal expression of nerve growth factor (NGF) was found in adenomyosis (AM). We collected AM foci from patients and eutopic endometrium from non-AM controls. Endometrium stromal cells (ESCs) were cultured. Different levels of 17β-estradiol, tumor necrosis factor (TNF), CoCl2, and H2O2 were added to the culture system separately, then the expression level of NGF in ESCs was detected. After adding different levels of NGF, the proliferation and apoptosis of ESCs and aromatase expression were detected. We found that 17β-estradiol promoted NGF production in AM ESCs but not in control ESCs; TNF promoted NGF production in both AM and control ESCs; and CoCl2 inhibited NGF production in control ESCs, but had no effect in AM ESCs. Nerve growth factor promoted the proliferation and synthesis of aromatase in AM ESCs. In conclusion, locally increased estrogen levels and inflammation may cause increased NGF production in the uterus of patients with AM. Nerve growth factor stimulated the proliferation and increased aromatase expression of ESCs from AM foci, suggesting NGF might contribute to the pathology and etiology of AM.

  14. Recombinant expression of human nerve growth factor beta in rabbit bone marrow mesenchymal stem cells.

    Science.gov (United States)

    Fan, Bo-Sheng; Lou, Ji-Yu

    2010-12-01

    Nerve growth factor (NGF) is required for the differentiation and maintenance of sympathetic and sensory neurons. In the present study, the recombinant expression of human nerve growth factor beta (hNGF-β) gene in rabbit bone marrow mesenchymal stem cells (rMSCs) was undertaken. Recombinant vector containing hNGF-β was constructed and transferred into rMSCs, the expressions of the exogenous in rMSCs were determined by reverse transcriptase PCR (RT-PCR), ELISA and Western blot, whereas the biological activity of recombinant hNGF-β was confirmed using PC12 cells and cultures of dorsal root ganglion neurons from chicken embryos. The results showed that the hNGF-β gene expressed successfully in the rMSCs, a polypeptide with a molecular weight of 13.2 kDa was detected. The maximal expression level of recombinant hNGF-β in rMSCs reached 126.8012 pg/10(6) cells, the mean concentration was 96.4473 pg/10(6) cells. The recombinant hNGF-β in the rMSCs showed full biological activity when compared to commercial recombinant hNGF-β.

  15. Pacemaker shift in the gastric antrum of guinea-pigs produced by excitatory vagal stimulation involves intramuscular interstitial cells.

    Science.gov (United States)

    Hirst, G D S; Dickens, E J; Edwards, F R

    2002-06-15

    Intracellular recordings were made from isolated bundles of the circular muscle layer of guinea-pig gastric antrum and the responses produced by stimulating intrinsic nerve fibres were examined. After abolishing the effects of stimulating inhibitory nerve terminals with apamin and L-nitroarginine (NOLA), transmural nerve stimulation often evoked a small amplitude excitatory junction potential (EJP) and invariably evoked a regenerative potential. Neurally evoked regenerative potentials had similar properties to those evoked in the same bundle by direct stimulation. EJPs and neurally evoked regenerative potentials were abolished by hyoscine suggesting that both resulted from the release of acetylcholine and activation of muscarinic receptors. Neurally evoked regenerative potentials, but not EJPs, were abolished by membrane hyperpolarization, caffeine and chloride channel blockers. In the intact antrum, excitatory vagal nerve stimulation increased the frequency of slow waves. Simultaneous intracellular recordings of pacemaker potentials from myenteric interstitial cells (ICC(MY)) and slow waves showed that the onset of each pacemaker potential normally preceded the onset of each slow wave but vagal stimulation caused the onset of each slow wave to precede each pacemaker potential. Together the observations suggest that during vagal stimulation there is a change in the origin of pacemaker activity with slow waves being initiated by intramuscular interstitial cells (ICC(IM)) rather than by ICC(MY).

  16. The use of fiber-reinforced scaffolds cocultured with Schwann cells and vascular endothelial cells to repair rabbit sciatic nerve defect with vascularization.

    Science.gov (United States)

    Gao, Hongyang; You, Yang; Zhang, Guoping; Zhao, Feng; Sha, Ziyi; Shen, Yong

    2013-01-01

    To explore the feasibility of biodegradable fiber-reinforced 3D scaffolds with satisfactory mechanical properties for the repair of long-distance sciatic nerve defect in rabbits and effects of vascularized graft in early stage on the recovery of neurological function, Schwann cells and vascular endothelial cells were cocultured in the fiber-reinforced 3D scaffolds. Experiment group which used prevascularized nerve complex for the repair of sciatic nerve defect and control group which only cultured with Schwann cells were set. The animals in both groups underwent electromyography to show the status of the neurological function recovery at 4, 8, and 16 weeks after the surgery. Sciatic nerve regeneration and myelination were observed under the light microscope and electron microscope. Myelin sheath thickness, axonal diameter, and number of myelinated nerve fiber were quantitatively analyzed using image analysis system. The recovery of foot ulcer, the velocity of nerve conduction, the number of regenerating nerve fiber, and the recovery of ultrastructure were increased in the experimental group than those in the control group. Prevascularized tissue engineered fiber-reinforced 3D scaffolds for the repair of sciatic nerve defects in rabbits can effectively promote the recovery of neurological function.

  17. The Use of Fiber-Reinforced Scaffolds Cocultured with Schwann Cells and Vascular Endothelial Cells to Repair Rabbit Sciatic Nerve Defect with Vascularization

    Directory of Open Access Journals (Sweden)

    Hongyang Gao

    2013-01-01

    Full Text Available To explore the feasibility of biodegradable fiber-reinforced 3D scaffolds with satisfactory mechanical properties for the repair of long-distance sciatic nerve defect in rabbits and effects of vascularized graft in early stage on the recovery of neurological function, Schwann cells and vascular endothelial cells were cocultured in the fiber-reinforced 3D scaffolds. Experiment group which used prevascularized nerve complex for the repair of sciatic nerve defect and control group which only cultured with Schwann cells were set. The animals in both groups underwent electromyography to show the status of the neurological function recovery at 4, 8, and 16 weeks after the surgery. Sciatic nerve regeneration and myelination were observed under the light microscope and electron microscope. Myelin sheath thickness, axonal diameter, and number of myelinated nerve fiber were quantitatively analyzed using image analysis system. The recovery of foot ulcer, the velocity of nerve conduction, the number of regenerating nerve fiber, and the recovery of ultrastructure were increased in the experimental group than those in the control group. Prevascularized tissue engineered fiber-reinforced 3D scaffolds for the repair of sciatic nerve defects in rabbits can effectively promote the recovery of neurological function.

  18. Role of sympathetic nerves in the establishment of metastatic breast cancer cells in bone

    Directory of Open Access Journals (Sweden)

    Florent Elefteriou

    2016-09-01

    Full Text Available The bone marrow microenvironment is characterized by its multicellular nature, and perhaps less obviously by the high mobility of multiple transient and stationary cell lineages present in this environment. The trafficking of hematopoietic and mesenchymal cells between the bone marrow and blood compartments is regulated by a number of bone marrow-derived factors. It is suspected that transformed metastatic cells “hijack” these processes to engraft into the skeleton and eventually cause the skeletal complications associated with metastatic disease. In this short review, experimental and association data supporting the contribution of a less recognized cell type of the bone marrow – the nerves of the sympathetic nervous system – to early events of the breast cancer bone metastatic process, are summarized.

  19. BNIP3 regulates AT101 [(--gossypol] induced death in malignant peripheral nerve sheath tumor cells.

    Directory of Open Access Journals (Sweden)

    Niroop Kaza

    Full Text Available Malignant peripheral nerve sheath tumors (MPNSTs are aggressive Schwann cell-derived sarcomas and are the leading cause of mortality in patients with neurofibromatosis type 1 (NF1. Current treatment modalities have been largely ineffective, resulting in a high rate of MPNST recurrence and poor five-year patient survival. This necessitates the exploration of alternative chemotherapeutic options for MPNST patients. This study sought to assess the cytotoxic effect of the BH3-mimetic AT101 [(--gossypol] on MPNST cells in vitro and to identify key regulators of AT101-induced MPNST cell death. We found that AT101 caused caspase-independent, non-apoptotic MPNST cell death, which was accompanied by autophagy and was mediated through HIF-1α induced expression of the atypical BH3-only protein BNIP3. These effects were mediated by intracellular iron chelation, a previously unreported mechanism of AT101 cytotoxicity.

  20. BNIP3 regulates AT101 [(-)-gossypol] induced death in malignant peripheral nerve sheath tumor cells.

    Science.gov (United States)

    Kaza, Niroop; Kohli, Latika; Graham, Christopher D; Klocke, Barbara J; Carroll, Steven L; Roth, Kevin A

    2014-01-01

    Malignant peripheral nerve sheath tumors (MPNSTs) are aggressive Schwann cell-derived sarcomas and are the leading cause of mortality in patients with neurofibromatosis type 1 (NF1). Current treatment modalities have been largely ineffective, resulting in a high rate of MPNST recurrence and poor five-year patient survival. This necessitates the exploration of alternative chemotherapeutic options for MPNST patients. This study sought to assess the cytotoxic effect of the BH3-mimetic AT101 [(-)-gossypol] on MPNST cells in vitro and to identify key regulators of AT101-induced MPNST cell death. We found that AT101 caused caspase-independent, non-apoptotic MPNST cell death, which was accompanied by autophagy and was mediated through HIF-1α induced expression of the atypical BH3-only protein BNIP3. These effects were mediated by intracellular iron chelation, a previously unreported mechanism of AT101 cytotoxicity.

  1. Facial Nerve Palsy: An Unusual Presenting Feature of Small Cell Lung Cancer

    Directory of Open Access Journals (Sweden)

    Ozcan Yildiz

    2011-01-01

    Full Text Available Lung cancer is the second most common type of cancer in the world and is the most common cause of cancer-related death in men and women; it is responsible for 1.3 million deaths annually worldwide. It can metastasize to any organ. The most common site of metastasis in the head and neck region is the brain; however, it can also metastasize to the oral cavity, gingiva, tongue, parotid gland and lymph nodes. This article reports a case of small cell lung cancer presenting with metastasis to the facial nerve.

  2. HLA-DR-expressing cells and T-lymphocytes in sural nerve biopsies

    DEFF Research Database (Denmark)

    Schrøder, H D; Olsson, T; Solders, G;

    1988-01-01

    was confirmed. HLA-DR expression was found in all biopsies and thus was not restricted to any particular type of neuropathy. The HLA-DR expression appeared to correlate with severity and activity of the neuropathy. HLA-DR-expressing macrophages wrapping myelinated fibers were prominent in primary demyelinating......Thirty-five sural nerve biopsies were stained immunohistochemically for HLA-DR antigen. HLA-DR was expressed on nonmyelinating Schwann cells, macrophages, vascular endothelium, and perineurium. By means of double immunofluorescence staining the identity of the HLA-DR presenting structures...

  3. Host cells and methods for producing isoprenyl alkanoates

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Taek Soon; Fortman, Jeffrey L.; Keasling, Jay D.

    2015-12-01

    The invention provides for a method of producing an isoprenyl alkanoate in a genetically modified host cell. In one embodiment, the method comprises culturing a genetically modified host cell which expresses an enzyme capable of catalyzing the esterification of an isoprenol and a straight-chain fatty acid, such as an alcohol acetyltransferase (AAT), wax ester synthase/diacylglycerol acyltransferase (WS/DGAT) or lipase, under a suitable condition so that the isoprenyl alkanoate is produced.

  4. Regulation of Axolotl (Ambystoma mexicanum Limb Blastema Cell Proliferation by Nerves and BMP2 in Organotypic Slice Culture.

    Directory of Open Access Journals (Sweden)

    Jeffrey Lehrberg

    Full Text Available We have modified and optimized the technique of organotypic slice culture in order to study the mechanisms regulating growth and pattern formation in regenerating axolotl limb blastemas. Blastema cells maintain many of the behaviors that are characteristic of blastemas in vivo when cultured as slices in vitro, including rates of proliferation that are comparable to what has been reported in vivo. Because the blastema slices can be cultured in basal medium without fetal bovine serum, it was possible to test the response of blastema cells to signaling molecules present in serum, as well as those produced by nerves. We also were able to investigate the response of blastema cells to experimentally regulated changes in BMP signaling. Blastema cells responded to all of these signals by increasing the rate of proliferation and the level of expression of the blastema marker gene, Prrx-1. The organotypic slice culture model provides the opportunity to identify and characterize the spatial and temporal co-regulation of pathways in order to induce and enhance a regenerative response.

  5. Regulation of Axolotl (Ambystoma mexicanum) Limb Blastema Cell Proliferation by Nerves and BMP2 in Organotypic Slice Culture.

    Science.gov (United States)

    Lehrberg, Jeffrey; Gardiner, David M

    2015-01-01

    We have modified and optimized the technique of organotypic slice culture in order to study the mechanisms regulating growth and pattern formation in regenerating axolotl limb blastemas. Blastema cells maintain many of the behaviors that are characteristic of blastemas in vivo when cultured as slices in vitro, including rates of proliferation that are comparable to what has been reported in vivo. Because the blastema slices can be cultured in basal medium without fetal bovine serum, it was possible to test the response of blastema cells to signaling molecules present in serum, as well as those produced by nerves. We also were able to investigate the response of blastema cells to experimentally regulated changes in BMP signaling. Blastema cells responded to all of these signals by increasing the rate of proliferation and the level of expression of the blastema marker gene, Prrx-1. The organotypic slice culture model provides the opportunity to identify and characterize the spatial and temporal co-regulation of pathways in order to induce and enhance a regenerative response.

  6. Novel three-dimensional nerve tissue engineering scaffolds and its biocompatibility with Schwann cells

    Institute of Scientific and Technical Information of China (English)

    YUAN Jian-dong; NIE Wen-bo; FU Qiang; LIAN Xiao-feng; HOU Tie-sheng; TAN Zhi-qing

    2009-01-01

    To develop a novel scaffolding method for the copolymers poly lactide-co-glycolide acid (PLGA) to construct a three-dimensional (3-D) scaffold and explore its biocompatibility through culturing Schwann cells (SCs) on it. Methods: The 3-D scaffolds were made by means of melt spinning, extension and weaving. The queueing disci-pline of the micro-channels were observed under a scan-ning electronic microscope (SEM).The sizes of the micropores and the factors of porosity were also measured. Sciatic nerves were harvested from 3-day-old Sprague Dawley (SD) rats for culture of SCs. SCs were separated, purified, and then implanted on PLGA scaffolds, gelatin sponge and poly-L-lysine (PLL)-coated tissue culture poly-styrene (TCPS) were used as biomaterial and cell-support-ive controls, respectively. The effect of PLGA on the adherence, proliferation and apoptosis of SCs were exam-ined in vitro in comparison with gelatin sponge and TCPS. Results: The micro-channels arrayed in parallel manners, and the pore sizes of the channels were uniform. No significant difference was found in the activity of Schwann cells cultured on PLEA and those on TCPS (P>0.05), and the DNA of PLGA scaffolds was not damaged. Conclusion: The 3-D scaffolds developed in this study have excellent structure and biocompatibility, which may be taken as a novel scaffold candidate for nerve-tissue engineering.

  7. Toxicity induced by cumene hydroperoxide in leech Retzius nerve cells: the protective role of glutathione.

    Science.gov (United States)

    Jovanovic, Zorica; Jovanovic, Svetlana

    2013-01-01

    In the present study, we studied the ability of glutathione (GSH) to detoxify exogenously applied cumene hydroperoxide (CHP). Exposure of leech Retzius nerve cells to CHP (1.5 mM) induced a marked prolongation of the spontaneous spike potential of these cells. Early after depolarization, and a cardiac-like action potential with a rapid depolarization followed by a sustained depolarization or plateau, which is terminated by a rapid repolarization were recorded. GSH (0.2 mM) significantly inhibited the effects of CHP on the duration of the action potential and suppressed CHP-induced spontaneous repetitive activity. Voltage-clamp recordings showed that CHP (1.5 mM) caused significant changes in the outward potassium currents. The fast and slow steady part of the potassium outward current was reduced by 46% and 39%, respectively. GSH applied in a concentration of 0.2 mM partially blocked the effect of CHP on the calcium-activated potassium currents. The fast and slow calcium-activated potassium currents were suppressed by about 20% and 15%, respectively. These results suggest that the neurotoxic effect of CHP on spontaneous spike electrogenesis and calcium-activated potassium currents of leech Retzius nerve cells was reduced in the presence of GSH.

  8. Evidence for a intimate relationship between mast cells and nerve fibers in the tongue of the frog, Rana esculenta

    Energy Technology Data Exchange (ETDEWEB)

    Chieffi Baccari, Gabriella; Minucci, Sergio [Naples, II Univ. (Italy). Dipt. di Fisiologia Umana e Funzioni Biologiche Integrate `Filippo Bottazzi`

    1997-12-31

    Morphological and ultrastructural association of mast cells and nerve fibers were studied in the tongue of the frog Rana esculenta. The number of mast cells in the tongue (253 {+-} 45 / mm{sup 2}) is far the highest of the frog tissue as far as people know. They are distributed throughout the connective tissue among the muscular fibers, near arterioles and venules but predominantly in close association and within the nerves. They are often embedded in the endoneurium within a nerve bundle near to myelinic or unmyelinic fibers and in membrane-to-membrane contact with axonlike processes. Just for the richness of mast cells, the tongue of the frog could represent an useful model to study the relationship between these cells and the peripheral nervous system.

  9. Relationship between interstitial cells of Cajal, fibroblast-like cells and inhibitory motor nerves in the internal anal sphincter.

    Science.gov (United States)

    Cobine, Caroline A; Hennig, Grant W; Kurahashi, Masaaki; Sanders, Kenton M; Ward, Sean M; Keef, Kathleen D

    2011-04-01

    Interstitial cells of Cajal (ICC) have been shown to participate in nitrergic neurotransmission in various regions of the gastrointestinal (GI) tract. Recently, fibroblast-like cells, which are positive for platelet-derived growth factor receptor α (PDGFRα(+)), have been suggested to participate additionally in inhibitory neurotransmission in the GI tract. The distribution of ICC and PDGFRα(+) cell populations and their relationship to inhibitory nerves within the mouse internal anal sphincter (IAS) are unknown. Immunohistochemical techniques and confocal microscopy were therefore used to examine the density and arrangement of ICC, PDGFRα(+) cells and neuronal nitric-oxide-synthase-positive (nNOS(+)) nerve fibers in the IAS of wild-type (WT) and W/W ( v ) mice. Of the total tissue volume within the IAS circular muscle layer, 18% consisted in highly branched PDGFRα(+) cells (PDGFRα(+)-IM). Other populations of PDGFRα(+) cells were observed within the submucosa and along the serosal and myenteric surfaces. Spindle-shaped intramuscular ICC (ICC-IM) were present in the WT mouse IAS but were largely absent from the W/W ( v ) IAS. The ICC-IM volume (5% of tissue volume) in the WT mouse IAS was significantly smaller than that of PDGFRα(+)-IM. Stellate-shaped submucosal ICC (ICC-SM) were observed in the WT and W/W ( v ) IAS. Minimum surface distance analysis revealed that nNOS(+) nerve fibers were closely aligned with both ICC-IM and PDGFRα(+)-IM. An even closer association was seen between ICC-IM and PDGFRα(+)-IM. Thus, a close morphological arrangement exists between inhibitory motor neurons, ICC-IM and PDGFRα(+)-IM suggesting that some functional interaction occurs between them contributing to inhibitory neurotransmission in the IAS.

  10. Ultrastructural and cytochemical identification of apoptotic cell death accompanying development of the fetal rat olfactory nerve layer.

    Science.gov (United States)

    Pellier, V; Saucier, D; Oestreicher, A B; Astic, L

    1996-07-01

    It has been previously shown that the embryonic olfactory nerve contains, in addition to glial ensheathing cells, a large population of differentiated neurons that migrate from the developing olfactory epithelium, in close association with the olfactory axon fascicles. The purpose of our study was to verify the hypothesis according to which a process of physiological cell death might be involved in the progressive disappearance of these migrating neurons that has been reported during late embryonic stages in several immunocytochemical studies. To do so, we have investigated the development of the olfactory nerve layer in rat embryos by using light and electron microscopy, with special reference to the presence of cell death processes within this structure. We have also applied the histochemical TUNEL method allowing in situ visualization of cells degenerating by apoptosis. In order to determine if neurons were present among dying cells, a procedure of double-labeling was performed by combining the DNA-specific bisbenzimide with two neuronal markers, the protein B-50/GAP-43 and the lectin Ulex europaeus I. Results brought out the precise temporal and spatial patterns of programmed cell death accompanying the morphogenesis of the olfactory nerve layer. A cell death process was observed within the olfactory nerve layer from its onset at embryonic day 13 (E13). While only few pycnotic cells were observed in E13 and E14 embryos, their number increased from E15 to reach a maximum at E16 and then diminished. Few dying cells were also observed along the olfactory axon fascicles when they penetrated the olfactory nerve layer. Degenerating cells appeared strongly TUNEL-labeled and exhibited morphological features of cell death by apoptosis. Double-labeling experiments revealed that some of the apoptotic cells were neurons. These observations indicate that apoptosis may account for the progressive decrease in the number of migrating neurons present within the embryonic

  11. Axonal regeneration and remyelination evaluation of chitosan/gelatin-based nerve guide combined with transforming growth factor-β1 and Schwann cells.

    Science.gov (United States)

    Nie, Xin; Deng, Manjing; Yang, Maojin; Liu, Luchuan; Zhang, Yongjie; Wen, Xiujie

    2014-01-01

    Despite efforts in peripheral nerve injury and regeneration, it is difficult to achieve a functional recovery following extended peripheral nerve lesions. Even if artificial nerve conduit, cell components and growth factors can enhance nerve regeneration, integration in peripheral nerve repair and regeneration remains yet to be explored. For this study, we used chitosan/gelatin nerve graft constructed with collagenous matrices as a vehicle for Schwann cells and transforming growth factor-β1 to bridge a 10-mm gap of the sciatic nerve and explored the feasibility of improving regeneration and reinnervation in rats. The nerve regeneration was assessed with functional recovery, electrophysiological test, retrograde labeling, and immunohistochemistry analysis during the post-operative period of 16 weeks. The results showed that the internal sides of the conduits were compact enough to prevent the connective tissues from ingrowth. Nerve conduction velocity, average regenerated myelin area, and myelinated axon count were similar to those treated with autograft (p > 0.05) but significantly higher than those bridged with chitosan/gelatin nerve graft alone (p regeneration and remyelination. A designed graft incorporating all of the tissue-engineering strategies for peripheral nerve regeneration may provide great progress in tissue engineering for nerve repair.

  12. Axonal outgrowth is associated with increased ERK 1/2 activation but decreased caspase 3 linked cell death in Schwann cells after immediate nerve repair in rats

    Directory of Open Access Journals (Sweden)

    Kanje Martin

    2011-01-01

    Full Text Available Abstract Background Extracellular-signal regulated kinase (ERK1/2 is activated by nerve damage and its activation precedes survival and proliferation of Schwann cells. In contrast, activation of caspase 3, a cysteine protease, is considered as a marker for apoptosis in Schwann cells. In the present study, axonal outgrowth, activation of ERK1/2 by phosphorylation (p-ERK 1/2 and immunoreactivity of cleaved caspase 3 were examined after immediate, delayed, or no repair of transected rat sciatic nerves. Results Axonal outgrowth, detected by neurofilament staining, was longer after immediate repair than after either the delayed or no repair conditions. Immediate repair also showed a higher expression of p-ERK 1/2 and a lower number of cleaved caspase 3 stained Schwann cells than after delayed nerve repair. If the transected nerve was not repaired a lower level of p-ERK 1/2 was found than in either the immediate or delayed repair conditions. Axonal outgrowth correlated to p-ERK 1/2, but not clearly with cleaved caspase 3. Contact with regenerating axons affected Schwann cells with respect to p-ERK 1/2 and cleaved caspase 3 after immediate nerve repair only. Conclusion The decreased regenerative capacity that has historically been observed after delayed nerve repair may be related to impaired activation of Schwann cells and increased Schwann cell death. Outgrowing axons influence ERK 1/2 activation and apoptosis of Schwann cells.

  13. A rapid and versatile method for the isolation, purification and cryogenic storage of Schwann cells from adult rodent nerves.

    Science.gov (United States)

    Andersen, Natalia D; Srinivas, Shruthi; Piñero, Gonzalo; Monje, Paula V

    2016-08-23

    We herein developed a protocol for the rapid procurement of adult nerve-derived Schwann cells (SCs) that was optimized to implement an immediate enzymatic dissociation of fresh nerve tissue while maintaining high cell viability, improving yields and minimizing fibroblast and myelin contamination. This protocol introduces: (1) an efficient method for enzymatic cell release immediately after removal of the epineurium and extensive teasing of the nerve fibers; (2) an adaptable drop-plating method for selective cell attachment, removal of myelin debris, and expansion of the initial SC population in chemically defined medium; (3) a magnetic-activated cell sorting purification protocol for rapid and effective fibroblast elimination; and (4) an optional step of cryopreservation for the storage of the excess of cells. Highly proliferative SC cultures devoid of myelin and fibroblast growth were obtained within three days of nerve processing. Characterization of the initial, expanded, and cryopreserved cell products confirmed maintenance of SC identity, viability and growth rates throughout the process. Most importantly, SCs retained their sensitivity to mitogens and potential for differentiation even after cryopreservation. To conclude, this easy-to-implement and clinically relevant protocol allows for the preparation of expandable homogeneous SC cultures while minimizing time, manipulation of the cells, and exposure to culture variables.

  14. Produced Water Treatment Using Microbial Fuel Cell Technology

    Energy Technology Data Exchange (ETDEWEB)

    Borole, A. P.; Campbell, R. [Campbell Applied Physics

    2011-05-20

    ORNL has developed a treatment for produced water using a combination of microbial fuel cells and electrosorption. A collaboration between Campbell Applied Physics and ORNL was initiated to further investigate development of the technology and apply it to treatment of field produced water. The project successfully demonstrated the potential of microbial fuel cells to generate electricity from organics in produced water. A steady voltage was continuously generated for several days using the system developed in this study. In addition to the extraction of electrical energy from the organic contaminants, use of the energy at the representative voltage was demonstrated for salts removal or desalination of the produced water. Thus, the technology has potential to remove organic as well as ionic contaminants with minimal energy input using this technology. This is a novel energy-efficient method to treat produced water. Funding to test the technology at larger scale is being pursued to enable application development.

  15. Human umbilical cord blood stem cells and brain-derived neurotrophic factor for optic nerve injury:a biomechanical evaluation

    Institute of Scientific and Technical Information of China (English)

    Zhong-jun Zhang; Ya-jun Li; Xiao-guang Liu; Feng-xiao Huang; Tie-jun Liu; Dong-mei Jiang; Xue-man Lv; Min Luo

    2015-01-01

    Treatment for optic nerve injury by brain-derived neurotrophic factor or the transplantation of human umbilical cord blood stem cells has gained progress, but analysis by biomechanical indicators is rare. Rabbit models of optic nerve injury were established by a clamp. At 7 days after injury, the vitreous body received a one-time injection of 50 μg brain-derived neurotrophic factor or 1 × 106 human umbilical cord blood stem cells. After 30 days, the maximum load, max-imum stress, maximum strain, elastic limit load, elastic limit stress, and elastic limit strain had clearly improved in rabbit models of optical nerve injury after treatment with brain-derived neu-rotrophic factor or human umbilical cord blood stem cells. The damage to the ultrastructure of the optic nerve had also been reduced. These ifndings suggest that human umbilical cord blood stem cells and brain-derived neurotrophic factor effectively repair the injured optical nerve, im-prove biomechanical properties, and contribute to the recovery after injury.

  16. 17β-Estradiol Promotes Schwann Cell Proliferation and Differentiation, Accelerating Early Remyelination in a Mouse Peripheral Nerve Injury Model

    Directory of Open Access Journals (Sweden)

    Yan Chen

    2016-01-01

    Full Text Available Estrogen induces oligodendrocyte remyelination in response to demyelination in the central nervous system. Our objective was to determine the effects of 17β-estradiol (E2 on Schwann cell function and peripheral nerve remyelination after injury. Adult male C57BL/6J mice were used to prepare the sciatic nerve transection injury model and were randomly categorized into control and E2 groups. To study myelination in vitro, dorsal root ganglion (DRG explant culture was prepared using 13.5-day-old mouse embryos. Primary Schwann cells were isolated from the sciatic nerves of 1- to 3-day-old Sprague–Dawley rats. Immunostaining for myelin basic protein (MBP expression and toluidine blue staining for myelin sheaths demonstrated that E2 treatment accelerates early remyelination in the “nerve bridge” region between the proximal and distal stumps of the transection injury site in the mouse sciatic nerve. The 5-bromo-2′-deoxyuridine incorporation assay revealed that E2 promotes Schwann cell proliferation in the bridge region and in the primary culture, which is blocked using AKT inhibitor MK2206. The in vitro myelination in the DRG explant culture determined showed that the MBP expression in the E2-treated group is higher than that in the control group. These results show that E2 promotes Schwann cell proliferation and myelination depending on AKT activation.

  17. Analysis of spatial relationships in three dimensions: tools for the study of nerve cell patterning

    Directory of Open Access Journals (Sweden)

    Raven Mary A

    2008-07-01

    Full Text Available Abstract Background Multiple technologies have been brought to bear on understanding the three-dimensional morphology of individual neurons and glia within the brain, but little progress has been made on understanding the rules controlling cellular patterning. We describe new matlab-based software tools, now available to the scientific community, permitting the calculation of spatial statistics associated with 3D point patterns. The analyses are largely derived from the Delaunay tessellation of the field, including the nearest neighbor and Voronoi domain analyses, and from the spatial autocorrelogram. Results Our tools enable the analysis of the spatial relationship between neurons within the central nervous system in 3D, and permit the modeling of these fields based on lattice-like simulations, and on simulations of minimal-distance spacing rules. Here we demonstrate the utility of our analysis methods to discriminate between two different simulated neuronal populations. Conclusion Together, these tools can be used to reveal the presence of nerve cell patterning and to model its foundation, in turn informing on the potential developmental mechanisms that govern its establishment. Furthermore, in conjunction with analyses of dendritic morphology, they can be used to determine the degree of dendritic coverage within a volume of tissue exhibited by mature nerve cells.

  18. Transcorneal electrical stimulation alters morphology and survival of retinal ganglion cells after optic nerve damage.

    Science.gov (United States)

    Henrich-Noack, Petra; Voigt, Nadine; Prilloff, Sylvia; Fedorov, Anton; Sabel, Bernhard A

    2013-05-24

    Traumatic optic nerve injury leads to retrograde death of retinal ganglion cells (RGCs), but transcorneal electrical stimulation (TES) can increase the cell survival rate. To understand the mechanisms and to further define the TES-induced effects we monitored in living animals RGC morphology and survival after optic nerve crush (ONC) in real time by using in vivo confocal neuroimaging (ICON) of the retina. ONC was performed in rats and ICON was performed before crush and on post-lesion days 3, 7 and 15 which allowed us to repeatedly record RGC number and size. TES or sham-stimulation were performed immediately after the crush and on post-injury day 11. Three days after ONC we detected a higher percentage of surviving RGCs in the TES group as compared to sham-treated controls. However, the difference was below significance level on day 7 and disappeared completely by day 15. The death rate was more variable amongst the TES-treated rats than in the control group. Morphological analysis revealed that average cell size changed significantly in the control group but not in stimulated animals and the morphological alterations of surviving neurons were smaller in TES-treated compared to control cells. In conclusion, TES delays post-traumatic cell death significantly. Moreover, we found "responder animals" which also benefited in the long-term from the treatment. Our in vivo cellular imaging results provide evidence that TES reduces ONC-associated neuronal swelling and shrinkage especially in RGCs which survived long-term. Further studies are now needed to determine the differences of responders vs. non-responders.

  19. Somatostatin-immunoreactive nerve cell bodies and fibers in the medulla oblongata et spinalis.

    Science.gov (United States)

    Forssmann, W G; Burnweit, C; Shehab, T; Triepel, J

    1979-10-01

    Complete serial sectioning of the medulla oblongata in monkey, cat, guinea pig, and japanese dancing mouse and incubation for somatostatin-immunoreaction was carried out. Numerous regions of the medulla oblongata such as the nucleus reticularis gigantocellularis, nucleus cuneatus et gracillis, nucleus raphe magnus, nucleus tractus solitarius, nucleus vestibularis, and parts of the oliva contain dense networks of somatostatin-immunoreactive nerve fibers. Cell bodies were seen in the nucleus reticularis medullae oblongatae. In the spinal cord the sections from each segment were analyzed, showing the highest concentrations of somatostatinergic fibers in the substantia gelantinosa of the columna dorsalis. Cell bodies were seen in the zona intermedia centralis, especially in the upper cervical segments. Many positive fibers were also seen in the entire zona intermedia and the columna ventralis. Especially prominent was the immunoreactivity in the zona intermediolateralis of the thoracic segments and the columna ventralis of the lower lumbar and sacral segments.

  20. Rita Levi-Montalcini and the discovery of NGF, the first nerve cell growth factor.

    Science.gov (United States)

    Aloe, Luigi

    2011-06-01

    The nerve growth factor (NGF) is a signaling protein, discovered by Rita Levi-Montalcini in the early 1950's for its effect on growth and differentiation of specific populations of neurons of the peripheral nervous system. Originally identified as neurite outgrowth-stimulating factor, later studies revealed that the purified molecule has a number of target cells in the central nervous system and on nonneuronal cells. Moreover, recent studies showed the potential therapeutic properties of NGF in neuropathies of the central and peripheral nervous system and diseases of the eye and skin. Here I briefly describe the discovery of NGF, the early studies of Rita LeviMontalcini, a pioneer in modern neuroscience, and my scientific and human experience working in her laboratory for over 40 years.

  1. Differentiation of mesenchymal stem cells into neuronal cells on fetal bovine acellular dermal matrix as a tissue engineered nerve scaffold

    Institute of Scientific and Technical Information of China (English)

    Yuping Feng; Jiao Wang; Shixin Ling; Zhuo Li; Mingsheng Li; Qiongyi Li; Zongren Ma; Sijiu Yu

    2014-01-01

    The purpose of this study was to assess fetal bovine acellular dermal matrix as a scaffold for supporting the differentiation of bone marrow mesenchymal stem cells into neural cells fol-lowing induction with neural differentiation medium. We performed long-term, continuous observation of cell morphology, growth, differentiation, and neuronal development using several microscopy techniques in conjunction with immunohistochemistry. We examined speciifc neu-ronal proteins and Nissl bodies involved in the differentiation process in order to determine the neuronal differentiation of bone marrow mesenchymal stem cells. The results show that bone marrow mesenchymal stem cells that differentiate on fetal bovine acellular dermal matrix display neuronal morphology with unipolar and bi/multipolar neurite elongations that express neuro-nal-speciifc proteins, includingβIII tubulin. The bone marrow mesenchymal stem cells grown on fetal bovine acellular dermal matrix and induced for long periods of time with neural differen-tiation medium differentiated into a multilayered neural network-like structure with long nerve ifbers that was composed of several parallel microifbers and neuronal cells, forming a complete neural circuit with dendrite-dendrite to axon-dendrite to dendrite-axon synapses. In addition, growth cones with filopodia were observed using scanning electron microscopy. Paraffin sec-tioning showed differentiated bone marrow mesenchymal stem cells with the typical features of neuronal phenotype, such as a large, round nucleus and a cytoplasm full of Nissl bodies. The data suggest that the biological scaffold fetal bovine acellular dermal matrix is capable of supporting human bone marrow mesenchymal stem cell differentiation into functional neurons and the subsequent formation of tissue engineered nerve.

  2. Interleukin 6 in intact and injured mouse peripheral nerves.

    Science.gov (United States)

    Reichert, F; Levitzky, R; Rotshenker, S

    1996-03-01

    The multifunctional cytokine interleukin 6 (IL-6) has direct growth, survival and differentiation effects on peripheral and central neurons. Furthermore, it can modulate the production by non-neuronal cells of other cytokines and growth factors, and thereby affect nerve cells indirectly. We have studied IL-6 expression and production in intact and injured peripheral nerves of C57/BL/6NHSD mice, which display the normal rapid progression of Wallerian degeneration. The IL-6 mRNA was detected in nerves degenerating in vitro or in vivo, but not in intact nerves. In vitro- and in vivo-degenerating nerve segments and neuroma nerve segments synthesized and secreted IL-6. The onset of IL-6 production was rapid and prolonged. It was detected as early as 2 h after injury and persisted for the entire period of 21 days tested after the injury. Of the non-neuronal cells that reside in intact and injured nerves, macrophages and fibroblasts were the major contributors to IL-6 production. We also studied IL-6 production in intact and injured nerves of mutant C57BL/6-WLD/OLA/NHSD mice, which display very slow progression of Wallerian degeneration. Injured nerves of C57BL/6-WLD/OLA/NHSD mice produced significantly lower amounts of IL-6 than did rapidly degenerating nerves of C57/BL/6NHSD mice.

  3. Endocannabinoids inhibit release of nerve growth factor by inflammation-activated mast cells.

    Science.gov (United States)

    Cantarella, Giuseppina; Scollo, Mimmo; Lempereur, Laurence; Saccani-Jotti, Gloria; Basile, Francesco; Bernardini, Renato

    2011-08-15

    Nerve growth factor (NGF) is a pleiotropic member of the neurotrophin family. Beside its neuronal effects, NGF plays a role in various processes, including angiogenesis. Mast cells release NGF and are among elements contributing to angiogenesis, a process regulated by arrays of factors, including the inhibitory cannabinoids. The possible inhibitory role of cannabinoids on mast cell-related NGF mitogenic effect on endothelial cells was then investigated. Human mastocytic cells HMC-1, challenged with PMA to yield release of NGF, were preincubated with the endocannabinoid PEA. Then, conditioned media were added to HUVEC cultures. PMA-activated HMC-1 cells released substantial amounts of NGF, whereas PEA inhibited PMA-induced NGF release. HUVEC proliferation increased after treatment with media from activated HMC-1 cells, while was reduced with media from HMC-1 cells treated with PEA. To characterize receptors mediating such effects of PEA, RT-PCR and western blot analysis were performed on HMC-1 cells. None of the two cannabinoid CB1 and CB2 receptors was expressed by HMC-1 cells, which on the other hand expressed the orphan receptor GPR55. PEA was ineffective in inhibiting NGF release from HMC-1 cells treated with PMA and transfected with positive GPR55 RNAi, whereas it induced significant reduction of NGF in cells transfected with the corresponding negative control RNAi. Results indicate that NGF released from inflammatory mast cells induces angiogenesis. Cannabinoids attenuate such pro-angiogenic effects of NGF. Finally, cannabinoids could be considered for antiangiogenic treatment in disorders characterized by prominent inflammation.

  4. Adenoviral-mediated glial cell line-derived neurotrophic factor gene transfer has a protective effect on sciatic nerve following constriction-induced spinal cord injury.

    Science.gov (United States)

    Chou, An-Kuo; Yang, Ming-Chang; Tsai, Hung-Pei; Chai, Chee-Yin; Tai, Ming-Hong; Kwan, Aij-Li; Hong, Yi-Ren

    2014-01-01

    Neuropathic pain due to peripheral nerve injury may be associated with abnormal central nerve activity. Glial cell-line-derived neurotrophic factor (GDNF) can help attenuate neuropathic pain in different animal models of nerve injury. However, whether GDNF can ameliorate neuropathic pain in the spinal cord dorsal horn (SCDH) in constriction-induced peripheral nerve injury remains unknown. We investigated the therapeutic effects of adenoviral-mediated GDNF on neuropathic pain behaviors, microglial activation, pro-inflammatory cytokine expression and programmed cell death in a chronic constriction injury (CCI) nerve injury animal model. In this study, neuropathic pain was produced by CCI on the ipsilateral SCDH. Mechanical allodynia was examined with von Frey filaments and thermal sensitivity was tested using a plantar test apparatus post-operatively. Target proteins GDNF-1, GDNFRa-1, MMP2, MMP9, p38, phospho-p38, ED1, IL6, IL1β, AIF, caspase-9, cleaved caspase-9, caspase-3, cleaved caspase-3, PARP, cleaved PARP, SPECTRIN, cleaved SPECTRIN, Beclin-1, PKCσ, PKCγ, iNOS, eNOS and nNOS were detected. Microglial activity was measured by observing changes in immunoreactivity with OX-42. NeuN and TUNEL staining were used to reveal whether apoptosis was attenuated by GDNF. Results showed that administrating GDNF began to attenuate both allodynia and thermal hyperalgesia at day 7. CCI-rats were found to have lower GDNF and GDNFRa-1 expression compared to controls, and GDNF re-activated their expression. Also, GDNF significantly down-regulated CCI-induced protein expression except for MMP2, eNOS and nNOS, indicating that the protective action of GDNF might be associated with anti-inflammation and prohibition of microglia activation. Immunocytochemistry staining showed that GDNF reduced CCI-induced neuronal apoptosis. In sum, GDNF enhanced the neurotrophic effect by inhibiting microglia activation and cytokine production via p38 and PKC signaling. GDNF could be a good

  5. Involvement of PACAP/ADNP signaling in the resistance to cell death in malignant peripheral nerve sheath tumor (MPNST) cells.

    Science.gov (United States)

    Castorina, Alessandro; Giunta, Salvatore; Scuderi, Soraya; D'Agata, Velia

    2012-11-01

    Malignant peripheral nerve sheath tumors (MPNSTs) are sarcomas able to grow under conditions of metabolic stress caused by insufficient nutrients or oxygen. Both pituitary adenylate cyclase-activating polypeptide (PACAP) and activity-dependent neuroprotective protein (ADNP) have glioprotective potential. However, whether PACAP/ADNP signaling is involved in the resistance to cell death in MPNST cells remains to be clarified. Here, we investigated the involvement of this signaling system in the survival response of MPNST cells against hydrogen peroxide (H(2)O(2))-evoked death both in the presence of normal serum (NS) and in serum-starved (SS) cells. Results showed that ADNP levels increased time-dependently (6-48 h) in SS cells. Treatment with PACAP38 (10(-9) to 10(-5) M) dose-dependently increased ADNP levels in NS but not in SS cells. PAC(1)/VPAC receptor antagonists completely suppressed PACAP-stimulated ADNP increase and partially reduced ADNP expression in SS cells. NS-cultured cells exposed to H(2)O(2) showed significantly reduced cell viability (~50 %), increased p53 and caspase-3, and DNA fragmentation, without affecting ADNP expression. Serum starvation significantly reduced H(2)O(2)-induced detrimental effects in MPNST cells, which were not further ameliorated by PACAP38. Altogether, these finding provide evidence for the involvement of an endogenous PACAP-mediated ADNP signaling system that increases MPNST cell resistance to H(2)O(2)-induced death upon serum starvation.

  6. Mast Cells Produce a Unique Chondroitin Sulfate Epitope.

    Science.gov (United States)

    Farrugia, Brooke L; Whitelock, John M; O'Grady, Robert; Caterson, Bruce; Lord, Megan S

    2016-02-01

    The granules of mast cells contain a myriad of mediators that are stored and protected by the sulfated glycosaminoglycan (GAG) chains that decorate proteoglycans. Whereas heparin is the GAG predominantly associated with mast cells, mast cell proteoglycans are also decorated with heparan sulfate and chondroitin sulfate (CS). This study investigated a unique CS structure produced by mast cells that was detected with the antibody clone 2B6 in the absence of chondroitinase ABC digestion. Mast cells in rodent tissue sections were characterized using toluidine blue, Leder stain and the presence of mast cell tryptase. The novel CS epitope was identified in rodent tissue sections and localized to cells that were morphologically similar to cells chemically identified as mast cells. The rodent mast cell-like line RBL-2H3 was also shown to express the novel CS epitope. This epitope co-localized with multiple CS proteoglycans in both rodent tissue and RBL-2H3 cultured cells. These findings suggest that the novel CS epitope that decorates mast cell proteoglycans may play a role in the way these chains are structured in mast cells.

  7. Human umbilical cord blood-derived mesenchymal stem cells promote regeneration of crush-injured rat sciatic nerves

    Institute of Scientific and Technical Information of China (English)

    Mi-Ae Sung; Jong-Ho Lee; Hun Jong Jung; Jung-Woo Lee; Jin-Yong Lee; Kang-Mi Pang; Sang Bae Yoo; Mohammad S. Alrashdan; Soung-Min Kim; Jeong Won Jahng

    2012-01-01

    Several studies have demonstrated that human umbilical cord blood-derived mesenchymal stem cells can promote neural regeneration following brain injury. However, the therapeutic effects of human umbilical cord blood-derived mesenchymal stem cells in guiding peripheral nerve regeneration remain poorly understood. This study was designed to investigate the effects of human umbilical cord blood-derived mesenchymal stem cells on neural regeneration using a rat sciatic nerve crush injury model. Human umbilical cord blood-derived mesenchymal stem cells (1 × 106) or a PBS control were injected into the crush-injured segment of the sciatic nerve. Four weeks after cell injection, brain-derived neurotrophic factor and tyrosine kinase receptor B mRNA expression at the lesion site was increased in comparison to control. Furthermore, sciatic function index, Fluoro Gold-labeled neuron counts and axon density were also significantly increased when compared with control. Our results indicate that human umbilical cord blood-derived mesenchymal stem cells promote the functional recovery of crush-injured sciatic nerves.

  8. Reconstruction of Multiple Facial Nerve Branches Using Skeletal Muscle-Derived Multipotent Stem Cell Sheet-Pellet Transplantation.

    Directory of Open Access Journals (Sweden)

    Kosuke Saito

    Full Text Available Head and neck cancer is often diagnosed at advanced stages, and surgical resection with wide margins is generally indicated, despite this treatment being associated with poor postoperative quality of life (QOL. We have previously reported on the therapeutic effects of skeletal muscle-derived multipotent stem cells (Sk-MSCs, which exert reconstitution capacity for muscle-nerve-blood vessel units. Recently, we further developed a 3D patch-transplantation system using Sk-MSC sheet-pellets. The aim of this study is the application of the 3D Sk-MSC transplantation system to the reconstitution of facial complex nerve-vascular networks after severe damage. Mouse experiments were performed for histological analysis and rats were used for functional examinations. The Sk-MSC sheet-pellets were prepared from GFP-Tg mice and SD rats, and were transplanted into the facial resection model (ST. Culture medium was transplanted as a control (NT. In the mouse experiment, facial-nerve-palsy (FNP scoring was performed weekly during the recovery period, and immunohistochemistry was used for the evaluation of histological recovery after 8 weeks. In rats, contractility of facial muscles was measured via electrical stimulation of facial nerves root, as the marker of total functional recovery at 8 weeks after transplantation. The ST-group showed significantly higher FNP (about three fold scores when compared to the NT-group after 2-8 weeks. Similarly, significant functional recovery of whisker movement muscles was confirmed in the ST-group at 8 weeks after transplantation. In addition, engrafted GFP+ cells formed complex branches of nerve-vascular networks, with differentiation into Schwann cells and perineurial/endoneurial cells, as well as vascular endothelial and smooth muscle cells. Thus, Sk-MSC sheet-pellet transplantation is potentially useful for functional reconstitution therapy of large defects in facial nerve-vascular networks.

  9. Renal erythropoietin-producing cells in health and disease

    Directory of Open Access Journals (Sweden)

    Tomokazu eSouma

    2015-06-01

    Full Text Available Erythropoietin (Epo is an indispensable erythropoietic hormone primarily produced from renal Epo-producing cells (REPs. Epo production in REPs is tightly regulated in a hypoxia-inducible manner to maintain tissue oxygen homeostasis. Insufficient Epo production by REPs causes renal anemia and anemia associated with chronic disorders. Recent studies have broadened our understanding of REPs from prototypic hypoxia-responsive cells to dynamic fibrogenic cells. In chronic kidney disease, REPs are the major source of scar-forming myofibroblasts and actively produce fibrogenic molecules, including inflammatory cytokines. Notably, myofibroblast-transformed REPs recover their original physiological properties after resolution of the disease insults, suggesting that renal anemia and fibrosis could be reversible to some extent. Therefore, understanding the plasticity of REPs will lead to the development of novel targeted therapeutics for both renal fibrosis and anemia. This review summarizes the regulatory mechanisms how hypoxia-inducible Epo gene expression is attained in health and disease conditions.

  10. Giant cell arteritis mimicking infiltrative leptomeningeal disease of the optic nerves.

    Science.gov (United States)

    Kornberg, Michael D; Ratchford, John N; Subramaniam, Rathan M; Probasco, John C

    2015-04-09

    A 67-year-old man presented with several days of progressive, painless left eye vision loss. He reported mild jaw claudication but denied headache, scalp tenderness or constitutional symptoms. Examination revealed palpable temporal arteries, blurring of the left optic disc, and 20/100 vision in the left eye with mild relative afferent pupillary defect. Inflammatory markers were sent, and methylprednisolone was initiated for presumptive giant cell arteritis (GCA). Erythrocyte sedimentation rate was normal, however, and C reactive protein was only mildly elevated, prompting further investigation. Orbital MRI revealed nodular enhancement of the optic nerve sheaths bilaterally from optic nerve head to chiasm, raising concern for an infiltrative leptomeningeal process such as sarcoidosis or lymphoma. Methylprednisolone was temporarily stopped while a broad work up for inflammatory and neoplastic causes was pursued. Fluorodeoxyglucose-positron emission tomography ultimately revealed hypermetabolism in the temporal, ophthalmic and occipital arteries suggesting GCA, which was confirmed by temporal artery biopsy. Steroids were restarted, and the patient's vision stabilised.

  11. Increased production of omega-3 fatty acids protects retinal ganglion cells after optic nerve injury in mice.

    Science.gov (United States)

    Peng, Shanshan; Shi, Zhe; Su, Huanxing; So, Kwok-Fai; Cui, Qi

    2016-07-01

    Injury to the central nervous system causes progressive degeneration of injured axons, leading to loss of the neuronal bodies. Neuronal survival after injury is a prerequisite for successful regeneration of injured axons. In this study, we investigated the effects of increased production of omega-3 fatty acids and elevation of cAMP on retinal ganglion cell (RGC) survival and axonal regeneration after optic nerve (ON) crush injury in adult mice. We found that increased production of omega-3 fatty acids in mice enhanced RGC survival, but not axonal regeneration, over a period of 3 weeks after ON injury. cAMP elevation promoted RGC survival in wild type mice, but no significant difference in cell survival was seen in mice over-producing omega-3 fatty acids and receiving intravitreal injections of CPT-cAMP, suggesting that cAMP elevation protects RGCs after injury but does not potentiate the actions of the omega-3 fatty acids. The observed omega-3 fatty acid-mediated neuroprotection is likely achieved partially through ERK1/2 signaling as inhibition of this pathway by PD98059 hindered, but did not completely block, RGC protection. Our study thus enhances our current understanding of neural repair after CNS injury, including the visual system.

  12. 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.

  13. Localization and biosynthesis of polyamines in insulin-producing cells

    DEFF Research Database (Denmark)

    Hougaard, D M; Larsson, L I; Nielsen, Jens Høiriis

    1986-01-01

    Two recently developed fluorescence cytochemical methods, specific for spermidine and spermine, were used to localize polyamines in the endocrine pancreas. The polyamines were restricted to the insulin-producing beta-cells and were mainly associated with the secretory granules. Chemical polyamine...

  14. A biosynthetic nerve guide conduit based on silk/SWNT/fibronectin nanocomposite for peripheral nerve regeneration.

    Science.gov (United States)

    Mottaghitalab, Fatemeh; Farokhi, Mehdi; Zaminy, Arash; Kokabi, Mehrdad; Soleimani, Masoud; Mirahmadi, Fereshteh; Shokrgozar, Mohammad Ali; Sadeghizadeh, Majid

    2013-01-01

    As a contribution to the functionality of nerve guide conduits (NGCs) in nerve tissue engineering, here we report a conduit processing technique through introduction and evaluation of topographical, physical and chemical cues. Porous structure of NGCs based on freeze-dried silk/single walled carbon nanotubes (SF/SWNTs) has shown a uniform chemical and physical structure with suitable electrical conductivity. Moreover, fibronectin (FN) containing nanofibers within the structure of SF/SWNT conduits produced through electrospinning process have shown aligned fashion with appropriate porosity and diameter. Moreover, fibronectin remained its bioactivity and influenced the adhesion and growth of U373 cell lines. The conduits were then implanted to 10 mm left sciatic nerve defects in rats. The histological assessment has shown that nerve regeneration has taken places in proximal region of implanted nerve after 5 weeks following surgery. Furthermore, nerve conduction velocities (NCV) and more myelinated axons were observed in SF/SWNT and SF/SWNT/FN groups after 5 weeks post implantation, indicating a functional recovery for the injured nerves. With immunohistochemistry, the higher S-100 expression of Schwann cells in SF/SWNT/FN conduits in comparison to other groups was confirmed. In conclusion, an oriented conduit of biocompatible SF/SWNT/FN has been fabricated with acceptable structure that is particularly applicable in nerve grafts.

  15. A biosynthetic nerve guide conduit based on silk/SWNT/fibronectin nanocomposite for peripheral nerve regeneration.

    Directory of Open Access Journals (Sweden)

    Fatemeh Mottaghitalab

    Full Text Available As a contribution to the functionality of nerve guide conduits (NGCs in nerve tissue engineering, here we report a conduit processing technique through introduction and evaluation of topographical, physical and chemical cues. Porous structure of NGCs based on freeze-dried silk/single walled carbon nanotubes (SF/SWNTs has shown a uniform chemical and physical structure with suitable electrical conductivity. Moreover, fibronectin (FN containing nanofibers within the structure of SF/SWNT conduits produced through electrospinning process have shown aligned fashion with appropriate porosity and diameter. Moreover, fibronectin remained its bioactivity and influenced the adhesion and growth of U373 cell lines. The conduits were then implanted to 10 mm left sciatic nerve defects in rats. The histological assessment has shown that nerve regeneration has taken places in proximal region of implanted nerve after 5 weeks following surgery. Furthermore, nerve conduction velocities (NCV and more myelinated axons were observed in SF/SWNT and SF/SWNT/FN groups after 5 weeks post implantation, indicating a functional recovery for the injured nerves. With immunohistochemistry, the higher S-100 expression of Schwann cells in SF/SWNT/FN conduits in comparison to other groups was confirmed. In conclusion, an oriented conduit of biocompatible SF/SWNT/FN has been fabricated with acceptable structure that is particularly applicable in nerve grafts.

  16. Chitosan-cross-linked nanofibrous PHBV nerve guide for rat sciatic nerve regeneration across a defect bridge.

    Science.gov (United States)

    Biazar, Esmaeil; Keshel, Saeed Heidari

    2013-01-01

    The aim of this study was to produce a chitosan-cross-linked nanofibrous biodegradable poly (3-hydroxybutyrate-co-3-hydroxyvalerate) nerve conduit. The artificial nerve scaffold designed by electrospinning method and cross-linked with chitosan by chemical method. Afterwards, the scaffolds were evaluated by microscopic, physical, and mechanical analyses and cell culture assays with Schwann cells. The conduits were implanted into a 10 mm gap in the sciatic nerves of the rats. Four months after surgery, the regenerated nerves were evaluated by macroscopic assessments and histology. This polymeric conduit had sufficiently good mechanical properties to serve as a nerve guide. Cellular experiments showed a better cell adhesion, growth, and proliferation inside the cross-linked nanofibrous scaffolds compared with un-cross-linked ones, also Schwann cells well attached on chitosan-cross-linked nanofibrous surface. The in vivo results demonstrated that in the nanofibrous graft, the sciatic nerve trunk had been reconstructed with restoration of nerve continuity and formatted nerve fibers with myelination. This neural conduit appears to have the right organization for testing in vivo nerve tissue engineering studies.

  17. Long-term delivery of nerve growth factor by encapsulated cell biodelivery in the Göttingen minipig basal forebrain

    DEFF Research Database (Denmark)

    Fjord-Larsen, Lone; Kusk, Philip; Tornøe, Jens;

    2010-01-01

    Nerve growth factor (NGF) prevents cholinergic degeneration in Alzheimer's disease (AD) and improves memory in AD animal models. In humans, the safe delivery of therapeutic doses of NGF is challenging. For clinical use, we have therefore developed an encapsulated cell (EC) biodelivery device...

  18. Expression patterns of cell cycle components in sporadic and neurofibromatosis type 1-related malignant peripheral nerve sheath tumors

    NARCIS (Netherlands)

    Agesen, Trude Holmeide; Florenes, Viva Ann; Molenaar, Willemina M.; Lind, Guro E.; Berner, Jeane-Marie; Plaat, Boudewijn E.C.; Komdeur, Rudy; Myklebost, Ola; van den Berg, Eva; Lothe, Ragnhild A.

    2005-01-01

    The molecular biology underlying the development of highly malignant peripheral nerve sheath tumors (MPNSTs) remains mostly unknown. In the present study, the expression pattern of 10 selected cell cycle components is investigated in a series of 15 MPNSTs from patients with (n = 9) or without (n = 5

  19. Positive effects of bFGF modified rat amniotic epithelial cells transplantation on transected rat optic nerve.

    Directory of Open Access Journals (Sweden)

    Jia-Xin Xie

    Full Text Available Effective therapy for visual loss caused by optic nerve injury or diseases has not been achieved even though the optic nerve has the regeneration potential after injury. This study was designed to modify amniotic epithelial cells (AECs with basic fibroblast growth factor (bFGF gene, preliminarily investigating its effect on transected optic nerve.A human bFGF gene segment was delivered into rat AECs (AECs/hbFGF by lentiviral vector, and the gene expression was examined by RT-PCR and ELISA. The AECs/hbFGF and untransfected rat AECs were transplanted into the transected site of the rat optic nerve. At 28 days post transplantation, the survival and migration of the transplanted cells was observed by tracking labeled cells; meanwhile retinal ganglion cells (RGCs were observed and counted by employing biotin dextran amine (BDA and Nissl staining. Furthermore, the expression of growth associated protein 43 (GAP-43 within the injury site was examined with immunohistochemical staining.The AECs/hbFGF was proven to express bFGF gene and secrete bFGF peptide. Both AECs/hbFGF and AECs could survive and migrate after transplantation. RGCs counting implicated that RGCs numbers of the cell transplantation groups were significantly higher than that of the control group, and the AECs/hbFGF group was significantly higher than that of the AECs group. Moreover GAP-43 integral optical density value in the control group was significantly lower than that of the cell transplantation groups, and the value in the AECs/hbFGF group was significantly higher than that of the AECs group.AECs modified with bFGF could reduce RGCs loss and promote expression of GAP-43 in the rat optic nerve transected model, facilitating the process of neural restoration following injury.

  20. Nerve growth factor protects against palmitic acid-induced injury in retinal ganglion cells

    Institute of Scientific and Technical Information of China (English)

    Pan-shi Yan; Shu Tang; Hai-feng Zhang; Yuan-yuan Guo; Zhi-wen Zeng; Qiang Wen

    2016-01-01

    Accumulating evidence supports an important role for nerve growth factor (NGF) in diabetic retinopathy. We hypothesized that NGF has a protective effect on rat retinal ganglion RGC-5 cells injured by palmitic acid (PA), a metabolic factor implicated in the development of dia-betes and its complications. Our results show that PA exposure caused apoptosis of RGC-5 cells, while NGF protected against PA insult in a concentration-dependent manner. Additionally, NGF signiifcantly attenuated the levels of reactive oxygen species (ROS) and malondialde-hyde (MDA) in RGC-5 cells. Pathway inhibitor tests showed that the protective effect of NGF was completely reversed by LY294002 (PI3K inhibitor), Akt VIII inhibitor, and PD98059 (ERK1/2 inhibitor). Western blot analysis revealed that NGF induced the phosphorylation of Akt/FoxO1 and ERK1/2 and reversed the PA-evoked reduction in the levels of these proteins. These results indicate that NGF protects RGC-5 cells against PA-induced injury through anti-oxidation and inhibition of apoptosis by modulation of the PI3K/Akt and ERK1/2 sig-naling pathways.

  1. Appropriate modulation of autophagy sensitizes malignant peripheral nerve sheath tumor cells to treatment with imatinib mesylate.

    Science.gov (United States)

    Okano, Munehiro; Sakata, Naoki; Ueda, Satoshi; Takemura, Tsukasa

    2014-04-01

    Malignant peripheral nerve sheath tumor (MPNST), very rare in childhood, is a highly aggressive soft-tissue tumor. We experienced a case of a 7-year-old boy with MPNST who was treated with imatinib mesylate (imatinib) after the identification of platelet-derived growth factor receptor expression in his tumor. We were unable to observe clinical benefits of imatinib in this patient. Therefore, cellular reactions of imatinib were investigated in vitro using 3 MPNST cell lines. Imatinib induced cytotoxicity in vitro with variable IC50 values (11.7 to >30 μM). Induction of apoptosis was not a pivotal mechanism in the inhibitory effects. We found that the treatment of MPNST cell lines with imatinib induced autophagy. Suppression of the initiation of autophagy by 3-methyladenine or small interfering RNA (siRNA) against beclin-1 attenuated the imatinib-mediated cytotoxicity. In contrast, blocking the formation of autophagosomes or the development of autolysosomes using siRNA against microtubule-associated protein light chain 3B, bafilomycin A1, chloroquine, or an MEK1/2 inhibitor (U0126) enhanced the imatinib-induced cytotoxicity in MPNST cells. Our data showed that the imatinib-mediated autophagy can function as a cytotoxic mechanism and that appropriate modulation of autophagy may sensitize MPNST cells to imatinib, which in turn may be a novel therapeutic strategy for MPNST.

  2. Preparation of PLLA/bpV(pic) microspheres and their effect on nerve cells.

    Science.gov (United States)

    Lin, Qiang; Chen, Hai-yun; Li, Hao-shen; Cai, Yang-ting

    2014-02-01

    In this study, we prepared PLLA/bpV(pic) microspheres, a bpV(pic) controlled release system and examined their ability to protect nerve cells and promote axonal growth. PLLA microspheres were prepared by employing the o/w single emulsification-evaporation technique. Neural stem cells and dorsal root ganglia were divided into 3 groups in terms of the treatment they received: a routine medium group (cultured in DMEM), a PLLA microsphere group (DMEM containing PLLA microspheres alone) and a PLLA/bpV(pic) group [DMEM containing PLLA/bpV(pic) microspheres]. The effects of PLLA/bpV(pic) microspheres were evaluated by the live-dead test and measurement of axonal length. Our results showed that PLLA/bpV(pic) granulation rate was (88.2±5.6)%; particle size was (16.8±3.1)%, drug loading was (4.05±0.3)%; encapsulation efficiency was (48.5±1.8)%. The release time lasted for 30 days. In PLLA/bpV(pic) microsphere group, the cell survival rate was (95.2 ±4.77)%, and the length of dorsal root ganglion (DRG) was 718±95 μm, which were all significantly greater than those in ordinary routine medium group and PLLA microsphere group. This preliminary test results showed the PLLA/bpV(pic) microspheres were successfully prepared and they could promote the survival and growth of neural cells in DRG.

  3. Isolation and characterization of renal erythropoietin-producing cells from genetically produced anemia mice.

    Directory of Open Access Journals (Sweden)

    Xiaoqing Pan

    Full Text Available Understanding the nature of renal erythropoietin-producing cells (REPs remains a central challenge for elucidating the mechanisms involved in hypoxia and/or anemia-induced erythropoietin (Epo production in adult mammals. Previous studies have shown that REPs are renal peritubular cells, but further details are lacking. Here, we describe an approach to isolate and characterize REPs. We bred mice bearing an Epo gene allele to which green fluorescent protein (GFP reporter cDNA was knocked-in (Epo(GFP with mice bearing an Epo gene allele lacking the 3' enhancer (Epo(Δ3'E. Mice harboring the mutant Epo(GFP/Δ3'E gene exhibited anemia (average Hematocrit 18% at 4 to 6 days after birth, and this perinatal anemia enabled us to identify and purify REPs based on GFP expression from the kidney. Light and confocal microscopy revealed that GFP immunostaining was confined to fibroblastic cells that reside in the peritubular interstitial space, confirming our previous observation in Epo-GFP transgenic reporter assays. Flow cytometry analyses revealed that the GFP fraction constitutes approximately 0.2% of the whole kidney cells and 63% of GFP-positive cells co-express CD73 (a marker for cortical fibroblasts and Epo-expressing cells in the kidney. Quantitative RT-PCR analyses confirmed that Epo expression was increased by approximately 100-fold in the purified population of REPs compared with that of the unsorted cells or CD73-positive fraction. Gene expression analyses showed enrichment of Hif2α and Hif3α mRNA in the purified population of REPs. The genetic approach described here provides a means to isolate a pure population of REPs, allowing the analysis of gene expression of a defined population of cells essential for Epo production in the kidney. This has provided evidence that positive regulation by HIF2α and negative regulation by HIF3α might be necessary for correct renal Epo induction.

  4. Comprehensive Profiling of GPCR Expression in Ghrelin-Producing Cells.

    Science.gov (United States)

    Koyama, Hiroyuki; Iwakura, Hiroshi; Dote, Katsuko; Bando, Mika; Hosoda, Hiroshi; Ariyasu, Hiroyuki; Kusakabe, Toru; Son, Choel; Hosoda, Kiminori; Akamizu, Takashi; Kangawa, Kenji; Nakao, Kazuwa

    2016-02-01

    To determine the comprehensive G protein-coupled receptor (GPCR) expression profile in ghrelin-producing cells and to elucidate the role of GPCR-mediated signaling in the regulation of ghrelin secretion, we determined GPCR expression profiles by RNA sequencing in the ghrelin-producing cell line MGN3-1 and analyzed the effects of ligands for highly expressed receptors on intracellular signaling and ghrelin secretion. Expression of selected GPCRs was confirmed in fluorescence-activated cell-sorted fluorescently tagged ghrelin-producing cells from ghrelin-promoter CreERT2/Rosa-CAG-LSL-ZsGreen1 mice. Expression levels of GPCRs previously suggested to regulate ghrelin secretion including adrenergic-β1 receptor, GPR81, oxytocin receptor, GPR120, and somatostatin receptor 2 were high in MGN3-1 cells. Consistent with previous reports, isoproterenol and oxytocin stimulated the Gs and Gq pathways, respectively, whereas lactate, palmitate, and somatostatin stimulated the Gi pathway, confirming the reliability of current assays. Among other highly expressed GPCRs, prostaglandin E receptor 4 agonist prostaglandin E2 significantly stimulated the Gs pathway and ghrelin secretion. Muscarine, the canonical agonist of cholinergic receptor muscarinic 4, stimulated both the Gq and Gi pathways. Although muscarine treatment alone did not affect ghrelin secretion, it did suppress forskolin-induced ghrelin secretion, suggesting that the cholinergic pathway may play a role in counterbalancing the stimulation of ghrelin by Gs (eg, by adrenaline). In addition, GPR142 ligand tryptophan stimulated ghrelin secretion. In conclusion, we determined the comprehensive expression profile of GPCRs in ghrelin-producing cells and identified two novel ghrelin regulators, prostaglandin E2 and tryptophan. These results will lead to a greater understanding of the physiology of ghrelin and facilitate the development of ghrelin-modulating drugs.

  5. Nerve biopsy

    Science.gov (United States)

    Biopsy - nerve ... A nerve biopsy is most often done on a nerve in the ankle, forearm, or along a rib. The health care ... feel a prick and a mild sting. The biopsy site may be sore for a few days ...

  6. The neuroscience of Helmholtz and the theories of Johannes Müller. Part 1: Nerve cell structure, vitalism, and the nerve impulse.

    Science.gov (United States)

    Finger, Stanley; Wade, Nicholas J

    2002-06-01

    Hermann Helmholtz made monumental contributions to the neural sciences in the second half of the nineteenth century. Among his earliest achievements were experiments that challenged vitalism, microscopic studies on the structure of the nerve cell and its processes, and the first reasonable estimates of the speed of nerve transmission based on physiological experiments. In this, the first of a two-part article, we review Helmholtz's early contributions in biographical context and with reference to Johannes Müller's own thoughts. We reveal how Johannes Müller, considered by many to be the greatest physiologist of the first half of the nineteenth century, helped to launch and shape Helmholtz's career. We also show that Helmholtz was only willing to accept some of his mentor's theories, even though he had great admiration for Müller. The point will be made that Helmholtz owed a great debt to Müller, but even from his student days in Berlin he was an independent thinker with his own agenda, and never his strict disciple.

  7. Schwann Cells Overexpressing FGF-2 Alone or Combined with Manual Stimulation Do Not Promote Functional Recovery after Facial Nerve Injury

    Directory of Open Access Journals (Sweden)

    Kirsten Haastert

    2009-01-01

    Full Text Available Purpose. To determine whether transplantation of Schwann cells (SCs overexpressing different isoforms of fibroblast growth factor 2 (FGF-2 combined with manual stimulation (MS of vibrissal muscles improves recovery after facial nerve transection in adult rat. Procedures. Transected facial nerves were entubulated with collagen alone or collagen plus naïve SCs or transfected SCs. Half of the rats received daily MS. Collateral branching was quantified from motoneuron counts after retrograde labeling from 3 facial nerve branches. Quality assessment of endplate reinnervation was combined with video-based vibrissal function analysis. Results. There was no difference in the extent of collateral axonal branching. The proportion of polyinnervated motor endplates for either naïve SCs or FGF-2 over-expressing SCs was identical. Postoperative MS also failed to improve recovery. Conclusions. Neither FGF-2 isoform changed the extent of collateral branching or polyinnervation of motor endplates; furthermore, this motoneuron response could not be overridden by MS.

  8. Establishment of a new bovine leukosis virus producing cell line.

    Science.gov (United States)

    Beier, D; Riebe, R; Blankenstein, P; Starick, E; Bondzio, A; Marquardt, O

    2004-11-01

    Due to the prevalence of different bovine leukosis virus (BLV) species in the cattle population in Europe, problems may arise in the serological diagnosis of BLV infections. In addition, earlier investigations demonstrated that contamination of the BLV antigen-producing cell culture systems by bovine viral diarrhea virus (BVDV) may give rise to misinterpretation of serological test results after BVDV vaccination of cattle. By co-cultivation of peripheral leukocytes of a BLV-infected cow with a permanent sheep kidney cell line, a new BLV-producing cell line named PO714 was established. This line carries a BLV provirus of the Belgian species and has been tested to be free of a variety of possibly contaminating viruses and mycoplasms. Investigations of a panel of well-characterised sera by agar gel immunodiffusion (AGID) and capture ELISA (cELISA) tests using antigen prepared from this new cell line in comparison with antigen of the well-known cell line FLK/BLV yielded comparable results. False positive results caused by BVDV cross-reactions could be eliminated when tests were carried out with antigen derived from the new cell line.

  9. Uterine autonomic nerve innervation plays a crucial role in regulating rat uterine mast cell functions during embryo implantation.

    Science.gov (United States)

    Yuan, Xue-Jun; Huang, Li-Bo; Qiao, Hui-Li; Deng, Ze-Pei; Fa, Jing-Jing

    2009-12-01

    To explore the potential mechanism of how uterine innervations would affect the uterine mast cell (MC) population and functions during the periimplantation. We herein first examined the consequence of uterine neurectomy on embryo implantation events. We observed that amputation of autonomic nerves innervating the uterus led to on-time implantation failure in rats. Exploiting MC culture and ELISA approaches, we then further analyzed the effect of neurectomy on cellular histamine levels and its release from uterine MCs, to elucidate the relation of the autonomic nerves and local cellular immunity in the uterine during early pregnancy. We observed that disconnection of autonomic nerve innervation significantly increased the population of uterine MCs. Most interestingly, these increased number of uterine MCs in neuroectomized rats contained a much reduced cellular level of histamine. Our subsequent challenge experiments revealed that uterine MCs in nerve amputated rats exhibited enhanced histamine releasing rate in response to substance P and antiIgE, suggesting loss of nerve innervation in the uterus not only increases the population of uterine MCs, but also facilitates the release of histamine from MCs, thus subsequently interfere with the normal implantation process. Collectively, our findings provide a new line of evidence supporting the concept that immune-neuro-endocrine network plays important role during pregnancy establishment and maintenance.

  10. Neuroprotective effects of BDNF and GDNF in intravitreally transplanted mesenchymal stem cells after optic nerve crush in mice

    Science.gov (United States)

    Hu, Zong-Li; Li, Ni; Wei, Xin; Tang, Li; Wang, Ting-Hua; Chen, Xiao-Ming

    2017-01-01

    AIM To assess the neuro-protective effect of bone marrow mesenchymal stem cells (BMSCs) on retinal ganglion cells (RGCs) following optic nerve crush in mice. METHODS C56BL/6J mice were treated with intravitreal injection of PBS, BMSCs, BDNF-interference BMSCs (BIM), and GDNF-interference BMSCs (GIM) following optic nerve crush, respectively. The number of surviving RGCs was determined by whole-mount retinas and frozen sections, while certain mRNA or protein was detected by q-PCR or ELISA, respectively. RESULTS The density (cell number/mm2) of RGCs was 410.77±56.70 in the retina 21d after optic nerve crush without any treatment, compared to 1351.39±195.97 in the normal control (PBIM (354.07+39.77) and GIM (326.67+33.37) than that without treatment (P<0.05). BMSCs injection improved the internal BDNF expression in retinas. CONCLUSION Optic nerve crush caused rust loss of RGCs and intravitreally transplanted BMSCs at some extent protected RGCs from death. The effect of BMSCs and level of BDNF in retinas are both related to BDNF and GDNF expression in BMSCs. PMID:28149774

  11. Stem Cell Ophthalmology Treatment Study (SCOTS for retinal and optic nerve diseases: a preliminary report

    Directory of Open Access Journals (Sweden)

    Jeffrey N Weiss

    2015-01-01

    Full Text Available In this report, we present the results of a single patient with optic neuropathy treated within the Stem Cell Ophthalmology Treatment Study (SCOTS. SCOTS is an Institutional Review Board approved clinical trial and is the largest ophthalmology stem cell study registered at the National Institutes of Health to date- www.clinicaltrials.gov Identifier NCT 01920867. SCOTS utilizes autologous bone marrow-derived stem cells in the treatment of optic nerve and retinal diseases. Pre- and post-treatment comprehensive eye exams were independently performed at the Wilmer Eye Institute at the Johns Hopkins Hospital, USA. A 27 year old female patient had lost vision approximately 5 years prior to enrollment in SCOTS. Pre-treatment best-corrected visual acuity at the Wilmer Eye Institute was 20/800 Right Eye (OD and 20/4,000 Left Eye (OS. Four months following treatment in SCOTS, the central visual acuity had improved to 20/100 OD and 20/40 OS.

  12. Effect of nerve regeneration factor on differentiation of PC12 cells and its signaling pathway

    Institute of Scientific and Technical Information of China (English)

    DING Fei; QIANG Liang; LIU Mei; GU Xingxing; GU Xiaosong

    2004-01-01

    The effects of nerve regeneration factor (NRF) on neuronal differentiation of PC12 cells and its signaling pathway are investigated by morphological observation and immunofluorescent cytochemical method, and the activity of ERK1/2 in NRF-treated PC12 cells in absence of serum is also studied by immuno-coprecipitation and Western blot analysis. The MEK1/2-specific inhibitor U0126, the broad-spectrum protein kinase C (PKC) inhibitor G6983 and tyrosine protein kinase (TPK) inhibitor genistein were used to determine the roles of the activation of ERK1/2 by NRF and the involvement of certain kinds of PKC or TPK receptor in this activation process. The results show that U0126 and G6983 inhibit the activation of ERK1/2 by NRF to different extents, while genistein has no effect on it, demonstrating that NRF remarkably induces neuronal differentiation of PC12 cells through activating ERK1/2 in a dose-dependent and time-dependent manner.

  13. Stem Cell Ophthalmology Treatment Study (SCOTS) for retinal and optic nerve diseases:a preliminary report

    Institute of Scientific and Technical Information of China (English)

    Jeffrey N Weiss; Steven Levy; Alexis Malkin

    2015-01-01

    In this report, we present the results of a single patient with optic neuropathy treated within the Stem Cell Ophthalmology Treatment Study (SCOTS). SCOTS is an Institutional Review Board approved clinical trial and is the largest ophthalmology stem cell study registered at the National Institutes of Health to date-www.clinicaltrials.gov Identifier NCT 01920867. SCOTS utilizes autologous bone marrow-derived stem cells in the treatment of optic nerve and retinal diseases. Pre-and post-treatment comprehensive eye exams were independently performed at the Wilmer Eye Institute at the Johns Hopkins Hospital, USA. A 27 year old female patient had lost vision approximately 5 years prior to enrollment in SCOTS. Pre-treatment best-corrected visual acuity at the Wilmer Eye Institute was 20/800 Right Eye (OD) and 20/4,000 Left Eye (OS). Four months following treatment in SCOTS, the central visual acuity had improved to 20/100 OD and 20/40 OS.

  14. Effect of a Nerve Graft Substitute Single-walled Carbon Nanotubes on Rat Pheochromocytoma Cells

    Institute of Scientific and Technical Information of China (English)

    WU Qian; ZHANG Juan; HU Zheqin

    2015-01-01

    Carbon nanotubes (CNTs) were extensively explored for their beneficial use in nervous system tissue engineering. However, an important concern regarding the use of CNTs is their toxicity during the interaction between cells and the nano particles. The rat pheochromocytoma cell line (PC12) was co-cultured with three types of single-walled carbon nanotubes (SWNTs), purified raw SWNTs (C), hydroxyl purified SWNTs (C-OH) and carboxyl purified SWNTs (C-COOH) at 25 µg/mL and 100 µg/ml. The experimental results revealed that SWNTs at the concentration below 100 µg/mL did not affect the cell viability. Notably, powerful antioxidant system in nerous system tissue is able to counteract with the toxicity of CNTs, which is characterized by the prominently enhanced expression of main antioxidant enzymes (catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx) and glutathione-S-transferase (GST)). Therefore, we believe that CNTs can be good candidates for the fabrication of biomedical scaffolds for the nerve tissue repair.

  15. Effect of vagus nerve stimulation on the secretory-granule volume of the principal cells of the mouse gallbladder epithelium.

    Science.gov (United States)

    Wahlin, T; Hulliger, M; Axelsson, H

    1979-07-01

    Experiments in mice were performed in order to investigate whether vagal activity could affect glycoprotein secretion from gallbladder principal cells. This cell type was studied with the electron microscope in control animals and after electric stimulation of the right or left nervus vagus. The volume density of glycoprotein containing granules was determined using morphometry. It was found that stimulation of the left vagus nerve significantly reduced the relative cellular volume of secretory granules in the principal cells of the gallbladder. Right vagus stimulation was accompanied by a weak but insignificant increase in secretory granule content. It is suggested that the left vagus nerve may exert a direct influence on glycoprotein secretion from gallbladder principal cells.

  16. Correction of surgically induced erectile dysfunction in the rat by repairing ablated cavernous nerve with neural stem cell grafts%神经干细胞移植重建勃起功能的研究

    Institute of Scientific and Technical Information of China (English)

    袁小旭; 邱剑光

    2010-01-01

    Objective To investigate the feasibility of restoring erectile function by repairing abla-ted cavernous nerve with neural stem cell grafts. Methods Forty-two male Sprague-Dawley rats (3-4 months old, weighing 300-400 g) were randomly divided into three groups: sham-operated controls (n = 14 ) subject to pelvic exploration without transection of the cavernous nerve; nerve ablation group (n = 14) having a 3 nun segment of cavernous nerve excised bilaterally; graft group (n = 14) having a 3 mm segment of cavernous nerve excised bilaterally, followed by immediate neural stem cell grafting. Two and 4 months postoperatively the rats of each group were electrostimulated to the cavernousal nerves to determine potency. Corpora cavernosa were harvested and nNOS-positive nerve fibers were examined by using immunohisto-chemistry. Results At the 2nd month postoperatively, electrical stimulation produced no erections in the nerve ablation or the graft group and 100 % erections in the sham-operated group. There was no significant difference in the nNOS-positive nerve fibers between the nerve ablation group and the graft group. At the 4th month postoperatively, 57. 14% of the grafted nerves produced erections upon electrical stimulation, significantly different from that in the nerve ablation group (7. 19% ). The nNOS-positive nerve fibers in the graft group (86. 0 ± 13.4) significantly differed from those in the nerve ablation group (26. 5±4. 3), and approximated the level of the sham-operated group (95.1 ± 7.7). Conclusion Neural stem cell graft-ing can be an effective method in restoring potency after injury.%目的 观察神经干细胞移植修复损伤的海绵体神经从而恢复勃起功能的可行性.方法 42只雄性SD大鼠(3~4个月龄和体质量300~400 g)随机分为假手术组、神经干细胞移植组和神经损伤组,每组14只.2、4个月后,海绵体神经电刺激检测大鼠阴茎勃起功能,免疫组织化学法检测海绵体内一氧化氮

  17. Biocompatibility evaluation of electrospun aligned poly(propylene carbonate) nanofibrous scaffolds with peripheral nerve tissues and cells in vitro

    Institute of Scientific and Technical Information of China (English)

    WANG Yu; ZHAO Zhe; ZHAO Bin; QI Hong-xu; PENG Jiang; ZHANG Li; XU Wen-jing; HU Ping; LU Shi-bi

    2011-01-01

    Background Peripheral nerve regeneration across large gaps is clinically challenging. Scaffold design plays a pivotal role in nerve tissue engineering. Recently, nanofibrous scaffolds have proven a suitable environment for cell attachment and proliferation due to similarities of their physical properties to natural extracellular matrix. Poly(propylene carbonate)(PPC) nanofibrous scaffolds have been investigated for vascular tissue engineering. However, no reports exist of PPC nanofibrous scaffolds for nerve tissue engineering. This study aimed to evaluate the potential role of aligned and random PPC nanofibrous scaffolds as substrates for peripheral nerve tissue and cells in nerve tissue engineering. Methods Aligned and random PPC nanofibrous scaffolds were fabricated by electrospinning and their chemical characterization were carried out using scanning electron microscopy (SEM). Dorsal root ganglia (DRG) from Sprague-Dawley rats were cultured on the nanofibrous substrates for 7 days. Neurite outgrowth and Schwann-cell migration from DRG were observed and quantified using immunocytochemistry and SEM. Schwann cells derived from rat sciatic nerves were cultured in electrospun PPC scaffold-extract fluid for 24, 48, 72 hours and 7 days. The viability of Schwann cells was evaluated by 3-[4,5-dimethyl(thiazol-2-yl)-2,5-diphenyl] tetrazolium bromide (MTT) assay. Results The diameter of aligned and random fibers ranged between 800 nm and 1200 nm, and the thickness of the films was approximately 10-20 μm. Quantification of aligned fiber films revealed approximately 90% alignment of all fibers along the longitudinal axis. However, with random fiber films, the alignment of fibers was random through all angle bins. Rat DRG explants were grown on PPC nanofiber films for up to 1 week. On the aligned fiber films, the majority of neurite outgrowth and Schwann cell migration from the DRG extended unidirectionally, parallel to the aligned fibers.However, on the random fiber films

  18. Migration and differentiation of neural progenitor cells after recurrent laryngeal nerve avulsion in rats.

    Directory of Open Access Journals (Sweden)

    Wan Zhao

    Full Text Available To investigate migration and differentiation of neural progenitor cells (NPCs from the ependymal layer to the nucleus ambiguus (NA after recurrent laryngeal nerve (RLN avulsion. All of the animals received a CM-DiI injection in the left lateral ventricle. Forty-five adult rats were subjected to a left RLN avulsion injury, and nine rats were used as controls. 5-Bromo-2-deoxyuridine (BrdU was injected intraperitoneally. Immunohistochemical analyses were performed in the brain stems at different time points after RLN injury. After RLN avulsion, the CM-DiI+ NPCs from the ependymal layer migrated to the lesioned NA. CM-DiI+/GFAP+ astrocytes, CM-DiI+/DCX+ neuroblasts and CM-DiI+/NeuN+ neurons were observed in the migratory stream. However, the ipsilateral NA included only CM-DiI+ astrocytes, not newborn neurons. After RLN avulsion, the NPCs in the ependymal layer of the 4th ventricle or central canal attempt to restore the damaged NA. We first confirm that the migratory stream includes both neurons and glia differentiated from the NPCs. However, only differentiated astrocytes are successfully incorporated into the NA. The presence of both cell types in the migratory process may play a role in repairing RLN injuries.

  19. Migration and differentiation of neural progenitor cells after recurrent laryngeal nerve avulsion in rats.

    Science.gov (United States)

    Zhao, Wan; Xu, Wen

    2014-01-01

    To investigate migration and differentiation of neural progenitor cells (NPCs) from the ependymal layer to the nucleus ambiguus (NA) after recurrent laryngeal nerve (RLN) avulsion. All of the animals received a CM-DiI injection in the left lateral ventricle. Forty-five adult rats were subjected to a left RLN avulsion injury, and nine rats were used as controls. 5-Bromo-2-deoxyuridine (BrdU) was injected intraperitoneally. Immunohistochemical analyses were performed in the brain stems at different time points after RLN injury. After RLN avulsion, the CM-DiI+ NPCs from the ependymal layer migrated to the lesioned NA. CM-DiI+/GFAP+ astrocytes, CM-DiI+/DCX+ neuroblasts and CM-DiI+/NeuN+ neurons were observed in the migratory stream. However, the ipsilateral NA included only CM-DiI+ astrocytes, not newborn neurons. After RLN avulsion, the NPCs in the ependymal layer of the 4th ventricle or central canal attempt to restore the damaged NA. We first confirm that the migratory stream includes both neurons and glia differentiated from the NPCs. However, only differentiated astrocytes are successfully incorporated into the NA. The presence of both cell types in the migratory process may play a role in repairing RLN injuries.

  20. Malignant peripheral nerve cell sheath tumour of the upper lip: a rare case

    Directory of Open Access Journals (Sweden)

    Joseph Ward

    2010-07-01

    Full Text Available We present the case of a malignant peripheral nerve sheath tumour (MPNST that developed on the upper lip of an 86 year old woman. MPNSTs are highly aggressive sarcomas that very rarely occur in the face. We know of no other reported cases of a malignant peripheral nerve sheath tumour arising from the upper lip.

  1. Allograft pretreatment for the repair of sciatic nerve defects:green tea polyphenolsversus radiation

    Institute of Scientific and Technical Information of China (English)

    Sheng-hu Zhou; Ping Zhen; Shen-song Li; Xiao-yan Liang; Ming-xuan Gao; Qi Tian; Xu-sheng Li

    2015-01-01

    Pretreatment of nerve allografts by exposure to irradiation or green tea polyphenols can elimi-nate neuroimmunogenicity, inhibit early immunological rejection, encourage nerve regeneration and functional recovery, improve tissue preservation, and minimize postoperative infection. In the present study, we investigate which intervention achieves better results. We produced a 1.0 cm sciatic nerve defect in rats, and divided the rats into four treatment groups: autograft, fresh nerve allograft, green tea polyphenol-pretreated (1 mg/mL, 4°C) nerve allograft, and irradiation-pre-treated nerve allograft (26.39 Gy/min for 12 hours; total 19 kGy). The animals were observed, and sciatic nerve electrophysiology, histology, and transmission electron microscopy were carried out at 6 and 12 weeks after grafting. The circumference and structure of the transplanted nerve in rats that received autografts or green tea polyphenol-pretreated nerve allografts were similar to those of the host sciatic nerve. Compared with the groups that received fresh or irradiation-pre-treated nerve allografts, motor nerve conduction velocity in the autograft and fresh nerve allograft groups was greater, more neurites grew into the allografts, Schwann cell proliferation was evident, and a large number of new blood vessels was observed; in addition, massive myelinated nerve ifbers formed, and abundant microiflaments and microtubules were present in the axoplasm. Our ifndings indicate that nerve allografts pretreated by green tea polyphenols are equivalent to trans-planting autologous nerves in the repair of sciatic nerve defects, and promote nerve regeneration. Pretreatment using green tea polyphenols is better than pretreatment with irradiation.

  2. 无细胞神经移植物复合骨髓间充质干细胞修复大鼠坐骨神经缺损%Repairing rat sciatic nerve defects with the compound of acellular nerve graft and bone marrow mesenchymal stem cells

    Institute of Scientific and Technical Information of China (English)

    张彩顺; 吕刚; 张基仁; 肖强

    2009-01-01

    BACKGROUND: Previous studies have successfully prepared the natural and biologically degraded acellular nerve graft and have proved the effect of promoting neural regeneration.OBJECTIVE: To construct tissue engineered artificial nerve with acellular nerve graft and bone marrow mesenchymal stem cells, and to observe the effect of promoting motor functional recovery and repairing rat sciatic nerve defects. DESIGN, TIME AND SETTING: Randomized control animal experiment was performed in the Medical TIssue Engineering Laboratory of the First Affiliated Hospital of Liaoning Medical University between June 2008 and February 2009. MATERIALS: Wistar adult healthy male rats weighing 180-200 g were used to prepare acellular nerve graft, while Wistar adult healthy male rats weighing 100-120 g were used to prepare bone marrow mesenchymal stem cells. Tissue engineered artificial nerve was produced with acellular nerve graft co-cultured with bone marrow mesenchymal stem cells. METHODS: Sixty Wistar adult healthy male rats weighing 180-200 g were induced sciatic nerve defect models, 15 mm long. SD rats were divided into three groups at random with 20 animals in each group. ①Experiment group: Rat sciatic nerve defects were bridged with tissue engineered artificial nerve. ②Blank control group: Rat sciatic nerve defects were bridged with tissue engineered nerve scaffold. ③Autologous nerve control group: Rat sciatic nerve defects were bridged with autologous nerve graft. MAIN OUTCOME MEASURES: At 12 weeks postoperation, the recovery of motor function was evaluated with gross observation, electrophysiology, histological observation and triceps surae wet weight.RESULTS: ①At 12 weeks postoperation, the toes at the operation side could separate and supported to the ground in the experiment group; there was no significant difference in the regenerated nerve conduction velocity between experimental group and autologous nerve graft group. ②At 12 weeks postoperation, histochemical

  3. Cdc42 Promotes Schwann Cell Proliferation and Migration Through Wnt/β-Catenin and p38 MAPK Signaling Pathway After Sciatic Nerve Injury.

    Science.gov (United States)

    Han, Bin; Zhao, Jun-Ying; Wang, Wu-Tao; Li, Zheng-Wei; He, Ai-Ping; Song, Xiao-Yang

    2017-01-17

    Schwann cells (SCs) are unique glial cells in the peripheral nerve and may secrete multiple neurotrophic factors, adhesion molecules, extracellular matrix molecules to form the microenvironment of peripheral nerve regeneration, guiding and supporting nerve proliferation and migration. Cdc42 plays an important regulatory role in dynamic changes of the cytoskeleton. However, there is a little study referred to regulation and mechanism of Cdc42 on glial cells after peripheral nerve injury. The present study investigated the role of Cdc42 in the proliferation and migration of SCs after sciatic nerve injury. Cdc42 expression was tested, showing that the mRNA and protein expression levels of Cdc42 were significantly up-regulated after sciatic nerve injury. Then, we isolated and purified SCs from injuried sciatic nerve at day 7. The purified SCs were transfected with Cdc42 siRNA and pcDNA3.1-Cdc42, and the cell proliferation, cell cycle and migration were assessed. The results implied that Cdc42 siRNA remarkably inhibited Schwann cell proliferation and migration, and resulted in S phase arrest. While pcDNA3.1-Cdc42 showed a contrary effect. Besides, we also observed that Cdc42 siRNA down-regulated the protein expression of β-catenin, Cyclin D1, c-myc and p-p38, which were up-regulated by pcDNA3.1-Cdc42. Meanwhile, the inhibitor of Wnt/β-catenin and p38 MAPK signaling pathway IWP-2 and SB203580 significantly inhibited the effect of pcDNA3.1-Cdc42 on cell proliferation and migration. Overall, our data indicate that Cdc42 regulates Schwann cell proliferation and migration through Wnt/β-catenin and p38 MAPK signaling pathway after sciatic nerve injury, which provides further insights into the therapy of the sciatic nerve injury.

  4. Trends in the design of nerve guidance channels in peripheral nerve tissue engineering.

    Science.gov (United States)

    Chiono, Valeria; Tonda-Turo, Chiara

    2015-08-01

    The current trend of peripheral nerve tissue engineering is the design of advanced nerve guidance channels (NGCs) acting as physical guidance for regeneration of nerves across lesions. NGCs should present multifunctional properties aiming to direct the sprouting of axons from the proximal nerve end, to concentrate growth factors secreted by the injured nerve ends, and to reduce the ingrowth of scar tissue into the injury site. A critical aspect in the design of NGCs is conferring them the ability to provide topographic, chemotactic and haptotactic cues that lead to functional nerve regeneration thus increasing the axon growth rate and avoiding or minimizing end-organ (e.g. muscle) atrophy. The present work reviews the recent state of the art in NGCs engineering and defines the external guide and internal fillers structural and compositional requirements that should be satisfied to improve nerve regeneration, especially in the case of large gaps (>2 cm). Techniques for NGCs fabrication were described highlighting the innovative approaches direct to enhance the regeneration of axon stumps compared to current clinical treatments. Furthermore, the possibility to apply stem cells as internal cues to the NGCs was discussed focusing on scaffold properties necessary to ensure cell survival. Finally, the optimized features for NGCs design were summarized showing as multifunctional cues are needed to produce NGCs having improved results in clinics.

  5. Analysis of the role of nerve growth factor in promoting cell survival during endoplasmic reticulum stress in PC12 cells.

    Science.gov (United States)

    Shimoke, Koji; Sasaya, Harue; Ikeuchi, Toshihiko

    2011-01-01

    Nerve growth factor (NGF) was first described by Rita Levi-Montalcini in the early 1960s from her studies of peripheral neurons. It has since been reported that NGF has the potential to elongate neurites or to prevent apoptosis via specific intracellular mechanisms. It has further been reported that as a component of these mechanisms, NGF binds to a specific receptor, TrkA, and thereby contributes to peripheral nerve cell functions or neuronal functions. It is noteworthy in this regard that pheochromocytoma 12 (PC12) cells express TrkA and respond to neurite outgrowth or anti-apoptotic signals by binding to NGF. Hence, PC12 cells have been used as an in vitro model system for the study of neuronal functions. It has been reported that endoplasmic reticulum (ER) stress is involved in neurodegenerative disorders, including Alzheimer's, Parkinson's, and Huntington's disease. The common link with regard to ER stress is that the neuronal cells die in these pathologies via specific intracellular mechanisms. This type of cell death, if it is apoptotic in nature, is termed ER stress-mediated apoptosis. In the process of ER stress-mediated apoptosis, the cleavage of pro-caspase-12 residing on the ER and the expression of glucose-regulated protein 78 (GRP78) can be observed. The expression of GRP78 protein is a characteristic of an unfolded protein response (UPR) via specific signal transduction pathways mediated by the unfolded protein response element (UPRE) in the upstream region of the grp78 gene so on. In ER stress-mediated apoptosis, a caspase cascade is also observed. To further clarify the mechanisms underlying ER stress-mediated apoptosis, a better understanding of the UPR is therefore important. In our current study, we describe a method for detecting gene induction via the UPR, focusing on GRP78 and caspase activities as the measurement end-points. The information generated by our method will accelerate our understanding of the pathophysiological processes leading

  6. Tumour Necrosis Factor α Enhances CCL2 and ICAM-1 Expression in Peripheral Nerve Microvascular Endoneurial Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Kelly A. Langert

    2013-01-01

    Full Text Available Recruitment and trafficking of autoreactive leucocytes across the BNB (blood–nerve barrier is an early pathological insult in GBS (Guillain-Barré syndrome, an aggressive autoimmune disorder of the PNS (peripheral nervous system. Whereas the aetiology and pathogenesis of GBS remain unclear, pro-inflammatory cytokines, including TNFα (tumour necrosis factor α, are reported to be elevated early in the course of GBS and may initiate nerve injury by activating the BNB. Previously, we reported that disrupting leucocyte trafficking in vivo therapeutically attenuates the course of an established animal model of GBS. Here, PNMECs (peripheral nerve microvascular endothelial cells that form the BNB were harvested from rat sciatic nerves, immortalized by SV40 (simian virus 40 large T antigen transduction and subsequently challenged with TNFα. Relative changes in CCL2 (chemokine ligand 2 and ICAM-1 (intercellular adhesion molecule 1 expression were determined. We report that TNFα elicits marked dose- and time-dependent increases in CCL2 and ICAM-1 mRNA and protein content and promotes secretion of functional CCL2 from immortalized and primary PNMEC cultures. TNFα-mediated secretion of CCL2 promotes, in vitro, the transendothelial migration of CCR2-expressing THP-1 monocytes. Increased CCL2 and ICAM-1 expression in response to TNFα may facilitate recruitment and trafficking of autoreactive leucocytes across the BNB in autoimmune disorders, including GBS.

  7. Nestin-expressing stem cells from the hair follicle can differentiate into motor neurons and reduce muscle atrophy after transplantation to injured nerves.

    Science.gov (United States)

    Liu, Fang; Zhang, Chuansen; Hoffman, Robert M

    2014-02-01

    We have previously shown that nestin-expressing hair follicle stem cells from the mouse and human are multipotent and can differentiate into many cell types, including neurons and glial cells. The nestin-expressing hair follicle stem cells can effect nerve and spinal cord repair upon transplantation in mouse models. In the present study, nestin-expressing hair follicle stem cells expressing red fluorescent protein (RFP) were induced by retinoic acid and fetal bovine serum to differentiate and then transplanted together with Matrigel into the transected distal sciatic or tibial nerve stump of transgenic nude mice ubiquitously expressing green fluorescent protein (GFP). Control mice were transplanted with Matrigel only. The transplanted cells appeared neuron like, with large round nuclei and long extensions. Immunofluorescence staining showed that some of the transplanted cells in the distal nerve stump expressed the neuron marker Tuj1 as well as motor neuron markers Isl 1/2 and EN1. These transplanted cells contacted each other as well as host nerve fibers. Two weeks post-transplantation, nerve fibers in the distal sciatic nerve stump of the transplanted mice had greater expression of motor neuron markers and neurotrophic factor-3 than those in the Matrigel-only transplanted mice. Muscle fiber areas in the nestin-expressing stem cell plus Matrigel-transplanted animals were much bigger than that in the Matrigel-only transplanted animals after 4 weeks. The present results suggest that transplanted nestin-expressing hair follicle stem cells can differentiate into motor neurons and reduce muscle atrophy after sciatic nerve transection. This study demonstrates a new and accessible neuron source to reduce muscle atrophy after nerve injury.

  8. Insulin-producing cells from embryonic stem cells rescues hyperglycemia via intra-spleen migration.

    Science.gov (United States)

    Ren, Meng; Shang, Changzhen; Zhong, Xiaomei; Guo, Ruomi; Lao, Guojuan; Wang, Xiaoyi; Cheng, Hua; Min, Jun; Yan, Li; Shen, Jun

    2014-12-23

    Implantation of embryonic stem cells (ESC)-derived insulin-producing cells has been extensively investigated for treatment of diabetes in animal models. However, the in vivo behavior and migration of transplanted cells in diabetic models remains unclear. Here we investigated the location and migration of insulin-producing cells labeled with superparamagnetic iron oxide (SPIO) using a dynamic MRI tracking method. SPIO labeled cells showed hypointense signal under the kidney subcapsules of diabetic mice on MRI, and faded gradually over the visiting time. However, new hypointense signal appeared in the spleen 1 week after transplantation, and became obvious with the time prolongation. Further histological examination proved the immigrated cells were insulin and C-peptide positive cells which were evenly distributed throughout the spleen. These intra-spleen insulin-producing cells maintained their protective effects against hyperglycemia in vivo, and these effects were reversed upon spleen removal. Transplantation of insulin-producing cells through spleen acquired an earlier blood glucose control as compared with that through kidney subcapsules. In summary, our data demonstrate that insulin-producing cells transplanted through kidney subcapsules were not located in situ but migrated into spleen, and rescues hyperglycemia in diabetic models. MRI may provide a novel tracking method for preclinical cell transplantation therapy of diabetes continuously and non-invasively.

  9. 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.

  10. Microencapsulation improves inhibitory effects of transplanted olfactory ensheathing cells on pain after sciatic nerve injury

    Directory of Open Access Journals (Sweden)

    Hao Zhao

    2015-01-01

    Full Text Available Olfactory bulb tissue transplantation inhibits P2X2/3 receptor-mediated neuropathic pain. However, the olfactory bulb has a complex cellular composition, and the mechanism underlying the action of purified transplanted olfactory ensheathing cells (OECs remains unclear. In the present study, we microencapsulated OECs in alginic acid, and transplanted free and microencapsulated OECs into the region surrounding the injured sciatic nerve in rat models of chronic constriction injury. We assessed mechanical nociception in the rat models 7 and 14 days after surgery by measuring paw withdrawal threshold, and examined P2X2/3 receptor expression in L 4-5 dorsal root ganglia using immunohistochemistry. Rats that received free and microencapsulated OEC transplants showed greater withdrawal thresholds than untreated model rats, and weaker P2X2/3 receptor immunoreactivity in dorsal root ganglia. At 14 days, paw withdrawal threshold was much higher in the microencapsulated OEC-treated animals. Our results confirm that microencapsulated OEC transplantation suppresses P2X2/3 receptor expression in L 4-5 dorsal root ganglia in rat models of neuropathic pain and reduces allodynia, and also suggest that transplantation of microencapsulated OECs is more effective than transplantation of free OECs for the treatment of neuropathic pain.

  11. Microencapsulation improves inhibitory effects of transplanted olfactory ensheathing cells on pain after sciatic nerve injury

    Institute of Scientific and Technical Information of China (English)

    Hao Zhao; Qing Li; Bao-lin Yang; Zeng-xu Liu; Qing Yu; Wen-jun Zhang; Keng Yuan; Hui-hong Zeng; Gao-chun Zhu; De-ming Liu

    2015-01-01

    Olfactory bulb tissue transplantation inhibits P2X2/3 receptor-mediated neuropathic pain. However, the olfactory bulb has a complex cellular composition, and the mechanism underlying the action of puriifed transplanted olfactory ensheathing cells (OECs) remains unclear. In the present study, we microencapsulated OECs in alginic acid, and transplanted free and microen-capsulated OECs into the region surrounding the injured sciatic nerve in rat models of chronic constriction injury. We assessed mechanical nociception in the rat models 7 and 14 days after surgery by measuring paw withdrawal threshold, and examined P2X2/3 receptor expression in L4–5 dorsal root ganglia using immunohistochemistry. Rats that received free and microencap-sulated OEC transplants showed greater withdrawal thresholds than untreated model rats, and weaker P2X2/3 receptor immunoreactivity in dorsal root ganglia. At 14 days, paw withdrawal threshold was much higher in the microencapsulated OEC-treated animals. Our results conifrm that microencapsulated OEC transplantation suppresses P2X2/3 receptor expression in L4–5 dorsal root ganglia in rat models of neuropathic pain and reduces allodynia, and also suggest that transplantation of microencapsulated OECs is more effective than transplantation of free OECs for the treatment of neuropathic pain.

  12. Toxicogenomic studies of human neural cells following exposure to organophosphorus chemical warfare nerve agent VX.

    Science.gov (United States)

    Gao, Xiugong; Lin, Hsiuling; Ray, Radharaman; Ray, Prabhati

    2013-05-01

    Organophosphorus (OP) compounds represent an important group of chemical warfare nerve agents that remains a significant and constant military and civilian threat. OP compounds are considered acting primarily via cholinergic pathways by binding irreversibly to acetylcholinesterase, an important regulator of the neurotransmitter acetylcholine. Many studies over the past years have suggested that other mechanisms of OP toxicity exist, which need to be unraveled by a comprehensive and systematic approach such as genome-wide gene expression analysis. Here we performed a microarray study in which cultured human neural cells were exposed to 0.1 or 10 μM of VX for 1 h. Global gene expression changes were analyzed 6, 24, and 72 h post exposure. Functional annotation and pathway analysis of the differentially expressed genes has revealed many genes, networks and canonical pathways that are related to nervous system development and function, or to neurodegenerative diseases such as Alzheimer's disease, Huntington's disease, and Parkinson's disease. In particular, the neuregulin pathway impacted by VX exposure has important implications in many nervous system diseases including schizophrenia. These results provide useful information valuable in developing suitable antidotes for more effective prevention and treatment of, as well as in developing biomarkers for, VX-induced chronic neurotoxicity.

  13. An analog VLSI implementation of the inner hair cell and auditory nerve using a dual AGC model.

    Science.gov (United States)

    Freedman, David S; Cohen, Howard I; Deligeorges, Socrates; Karl, Christian; Hubbard, Allyn E

    2014-04-01

    An analog inner hair cell and auditory nerve circuit using a dual AGC model has been implemented using 0.35 micron mixed-signal technology. A fully-differential current-mode architecture is used and the ability to correct channel mismatch is evaluated with matched layouts as well as with digital current tuning. The Meddis test paradigm is used to examine the analog implementation's auditory processing capabilities and investigate the circuit's ability to correct DC mismatch. The correction techniques used demonstrate the analog inner hair cell and auditory nerve circuit's potential use in low-power, multiple-sensor analog biomimetic systems with highly reproducible signal processing blocks on a single massively parallel integrated circuit.

  14. Dorsal root ganglion-derived Schwann cells combined with poly(lactic-co-glycolic acid)/chitosan conduits for the repair of sciatic nerve defects in rats.

    Science.gov (United States)

    Zhao, Li; Qu, Wei; Wu, Yuxuan; Ma, Hao; Jiang, Huajun

    2014-11-15

    Schwann cells, nerve regeneration promoters in peripheral nerve tissue engineering, can be used to repair both the peripheral and central nervous systems. However, isolation and purification of Schwann cells are complicated by contamination with fibroblasts. Current reported measures are mainly limited by either high cost or complicated procedures with low cell yields or purity. In this study, we collected dorsal root ganglia from neonatal rats from which we obtained highly purified Schwann cells using serum-free melanocyte culture medium. The purity of Schwann cells (> 95%) using our method was higher than that using standard medium containing fetal bovine serum. The obtained Schwann cells were implanted into poly(lactic-co-glycolic acid)/chitosan conduits to repair 10-mm sciatic nerve defects in rats. Results showed that axonal diameter and area were significantly increased and motor functions were obviously improved in the rat sciatic nerve tissue. Experimental findings suggest that serum-free melanocyte culture medium is conducive to purify Schwann cells and poly(lactic-co-glycolic acid)/chitosan nerve conduits combined with Schwann cells contribute to restore sciatic nerve defects.

  15. Dorsal root ganglion-derived Schwann cells combined with poly(lactic-co-glycolic acid)/chitosan conduits for the repair of sciatic nerve defects in rats

    Institute of Scientific and Technical Information of China (English)

    Li Zhao; Wei Qu; Yuxuan Wu; Hao Ma; Huajun Jiang

    2014-01-01

    Schwann cells, nerve regeneration promoters in peripheral nerve tissue engineering, can be used to repair both the peripheral and central nervous systems. However, isolation and puriifcation of Schwann cells are complicated by contamination with ifbroblasts. Current reported measures are mainly limited by either high cost or complicated procedures with low cell yields or purity. In this study, we collected dorsal root ganglia from neonatal rats from which we obtained highly puriifed Schwann cells using serum-free melanocyte culture medium. The purity of Schwann cells (> 95%) using our method was higher than that using standard medium containing fetal bovine serum. The obtained Schwann cells were implanted into poly(lactic-co-glycolic acid)/chi-tosan conduits to repair 10-mm sciatic nerve defects in rats. Results showed that axonal diameter and area were signiifcantly increased and motor functions were obviously improved in the rat sciatic nerve tissue. Experimental ifndings suggest that serum-free melanocyte culture medium is conducive to purify Schwann cells and poly(lactic-co-glycolic acid)/chitosan nerve conduits combined with Schwann cells contribute to restore sciatic nerve defects.

  16. Effects of collagen membranes enriched with in vitro-differentiated N1E-115 cells on rat sciatic nerve regeneration after end-to-end repair

    Directory of Open Access Journals (Sweden)

    Fornaro Michele

    2010-02-01

    Full Text Available Abstract Peripheral nerves possess the capacity of self-regeneration after traumatic injury but the extent of regeneration is often poor and may benefit from exogenous factors that enhance growth. The use of cellular systems is a rational approach for delivering neurotrophic factors at the nerve lesion site, and in the present study we investigated the effects of enwrapping the site of end-to-end rat sciatic nerve repair with an equine type III collagen membrane enriched or not with N1E-115 pre-differentiated neural cells. After neurotmesis, the sciatic nerve was repaired by end-to-end suture (End-to-End group, end-to-end suture enwrapped with an equine collagen type III membrane (End-to-EndMemb group; and end-to-end suture enwrapped with an equine collagen type III membrane previously covered with neural cells pre-differentiated in vitro from N1E-115 cells (End-to-EndMembCell group. Along the postoperative, motor and sensory functional recovery was evaluated using extensor postural thrust (EPT, withdrawal reflex latency (WRL and ankle kinematics. After 20 weeks animals were sacrificed and the repaired sciatic nerves were processed for histological and stereological analysis. Results showed that enwrapment of the rapair site with a collagen membrane, with or without neural cell enrichment, did not lead to any significant improvement in most of functional and stereological predictors of nerve regeneration that we have assessed, with the exception of EPT which recovered significantly better after neural cell enriched membrane employment. It can thus be concluded that this particular type of nerve tissue engineering approach has very limited effects on nerve regeneration after sciatic end-to-end nerve reconstruction in the rat.

  17. Human B cells produce chemokine CXCL10 in the presence of Mycobacterium tuberculosis specific T cells

    DEFF Research Database (Denmark)

    Hoff, Soren T; Salman, Ahmed M; Ruhwald, Morten

    2015-01-01

    BACKGROUND: The role of B cells in human host response to Mycobacterium tuberculosis (Mtb) infection is still controversial, but recent evidence suggest that B cell follicle like structures within the lung may influence host responses through regulation of the local cytokine environment....... A candidate for such regulation could be the chemokine CXCL10. CXCL10 is mainly produced by human monocytes, but a few reports have also found CXCL10 production by human B cells. The objective of this study was to investigate CXCL10 production by human B cells in response to in vitro stimulation with Mtb...... antigens. METHODOLOGY/PRINCIPAL FINDINGS: We analyzed human blood samples from 30 volunteer donors using multiparameter flow cytometry, and identified a subgroup of B cells producing CXCL10 in response to in vitro stimulation with antigens. T cells did not produce CXCL10, but CXCL10 production by B cells...

  18. Differentiation of dermis-derived multipotent cells into insulin-producing pancreatic cells in vitro

    Institute of Scientific and Technical Information of China (English)

    Chun-Meng Shi; Tian-Min Cheng

    2004-01-01

    AIM: To observe the plasticity of whether dermis-derived multipotent cells to differentiate into insulin-producing pancreatic cells in vitro.METHODS: A donal population of dermis-derived multipotent stem cells (DMCs) from newborn rat with the capacity to produce osteocytes, chondrocytes, adipocytes and neurons was used. The gene expression of cultured DMCs was assessed by DNA microarray using rat RGU34A gene expression probe arrays. DMCs were further cultured in the presence of insulin complex components (Insulintransferrin-selenium, ITS) to observe whether DMCs could be induced into insulin-producing pancreatic cells in vitro.RESULTS: DNA microarray analysis showed that cultured DMCs simultaneously expressed several genes associated with pancreatic cell, neural cell, epithelial cell and hepatocyte,widening its transcriptomic repertoire. When cultured in the specific induction medium containing ITS for pancreatic cells, DMCs differentiated into epithelioid cells that were positive for insulin detected by immunohistochemistry.CONCLUSION: Our data indicate that dermal multipotent cells may serve as a source of stem/progenitor cells for insulin-producing pancreatic cells.

  19. Tumors of the optic nerve

    DEFF Research Database (Denmark)

    Lindegaard, Jens; Heegaard, Steffen

    2009-01-01

    A variety of lesions may involve the optic nerve. Mainly, these lesions are inflammatory or vascular lesions that rarely necessitate surgery but may induce significant visual morbidity. Orbital tumors may induce proptosis, visual loss, relative afferent pupillary defect, disc edema and optic...... atrophy, but less than one-tenth of these tumors are confined to the optic nerve or its sheaths. No signs or symptoms are pathognomonic for tumors of the optic nerve. The tumors of the optic nerve may originate from the optic nerve itself (primary tumors) as a proliferation of cells normally present...... in the nerve (e.g., astrocytes and meningothelial cells). The optic nerve may also be invaded from tumors originating elsewhere (secondary tumors), invading the nerve from adjacent structures (e.g., choroidal melanoma and retinoblastoma) or from distant sites (e.g., lymphocytic infiltration and distant...

  20. Neurofibromin specific antibody differentiates malignant peripheral nerve sheath tumors (MPNST) from other spindle cell neoplasms.

    Science.gov (United States)

    Reuss, David E; Habel, Antje; Hagenlocher, Christian; Mucha, Jana; Ackermann, Ulrike; Tessmer, Claudia; Meyer, Jochen; Capper, David; Moldenhauer, Gerhard; Mautner, Victor; Frappart, Pierre-Olivier; Schittenhelm, Jens; Hartmann, Christian; Hagel, Christian; Katenkamp, Kathrin; Petersen, Iver; Mechtersheimer, Gunhild; von Deimling, Andreas

    2014-04-01

    Malignant peripheral nerve sheath tumors (MPNST) derive from the Schwann cell or perineurial cell lineage and occur either sporadically or in association with the tumor syndrome neurofibromatosis type 1 (NF1). MPNST often pose a diagnostic challenge due to their frequent lack of pathognomonic morphological or immunohistochemical features. Mutations in the NF1 tumor suppressor gene are found in all NF1-associated and many sporadic MPNST. The presence of NF1 mutation may have the potential to differentiate MPNST from several morphologically similar neoplasms; however, mutation detection is hampered by the size of the gene and the lack of mutational hot spots. Here we describe a newly developed monoclonal antibody binding to the C-terminus of neurofibromin (clone NFC) which was selected for optimal performance in routinely processed formalin-fixed and paraffin-embedded tissue. NFC immunohistochemistry revealed loss of neurofibromin in 22/25 (88 %) of NF1-associated and 26/61 (43 %) of sporadic MPNST. There was a strong association of neurofibromin loss with deletions affecting the NF1 gene (P < 0.01). In a series of 256 soft tissue tumors of different histotypes NFC staining showed loss of neurofibromin in 2/8 myxofibrosarcomas, 2/12 (16 %) pleomorphic liposarcomas, 1/16 (6 %) leiomyosarcomas, and 4/28 (14 %) unclassified undifferentiated pleomorphic sarcomas. However, loss of neurofibromin was not observed in 22 synovial sarcomas, 27 schwannomas, 23 solitary fibrous tumors, 14 low-grade fibromyxoid sarcomas, 50 dedifferentiated liposarcomas, 27 myxoid liposarcomas, 13 angiosarcomas, 9 extraskeletal myxoid chondrosarcomas, and 7 epitheloid sarcomas. Immunohistochemistry using antibody NFC may substantially facilitate sarcoma research and diagnostics.

  1. Microbes produce nanobacteria-like structures, avoiding cell entombment

    Science.gov (United States)

    Bontognali, Tomaso R. R.; Vasconcelos, Crisógono; Warthmann, Rolf J.; Dupraz, Christophe; Bernasconi, Stefano M.; McKenzie, Judith A.

    2008-08-01

    Microsedimentary structures referred to as nanobacteria-likeparticles were described from modern carbonate environments,where they form in close spatial association with sulfate-reducingbacteria (SRB). However, the exact mechanism of their formation,as well as their paleontological significance, remains controversial.Here we report on an investigation of microbe-mineral interactionsin experimentally produced carbonate globules. The experimentswere carried out under anoxic conditions at 30 °C with Desulfovibriobrasiliensis, a SRB known to mediate dolomite formation. Weobserved that extracellular polymeric substances (EPS) secretedby the microbial community play a key role in the mineralizationprocess. Nanobacteria-like particles represent the early stageof carbonate nucleation within the EPS, which progressivelyevolve to larger globules displaying a grainy texture. We excludedthe possibilities that these structures are fossils of nanobacteria,dissolution surfaces, or artifacts created during sample preparation.D. brasiliensis cells are predominantly located outside of theEPS aggregates where mineral growth takes place. As a result,they remain mobile and are rarely entombed within the mineral.This self-preservation behavior may not be limited to D. brasiliensis.Other microbes may produce, or may have produced during thegeological past, biogenic minerals through a similar process.Mineralization within EPS explains why microbial relics arenot necessarily present in biogenic carbonates.

  2. Feasibility of Human Amniotic Fluid Derived Stem Cells in Alleviation of Neuropathic Pain in Chronic Constrictive Injury Nerve Model.

    Directory of Open Access Journals (Sweden)

    Chien-Yi Chiang

    Full Text Available The neurobehavior of neuropathic pain by chronic constriction injury (CCI of sciatic nerve is very similar to that in humans, and it is accompanied by a profound local inflammation response. In this study, we assess the potentiality of human amniotic fluid derived mesenchymal stem cells (hAFMSCs for alleviating the neuropathic pain in a chronic constriction nerve injury model.This neuropathic pain animal model was conducted by four 3-0 chromic gut ligatures loosely ligated around the left sciatic nerve in Sprague-Dawley rats. The intravenous administration of hAFMSCs with 5x105 cells was conducted for three consecutive days.The expression IL-1β, TNF-α and synaptophysin in dorsal root ganglion cell culture was remarkably attenuated when co-cultured with hAFMSCs. The significant decrease of PGP 9.5 in the skin after CCI was restored by administration of hAFMSCs. Remarkably increased expression of CD 68 and TNF-α and decreased S-100 and neurofilament expression in injured nerve were rescued by hAFMSCs administration. Increases in synaptophysin and TNF-α over the dorsal root ganglion were attenuated by hAFMSCs. Significant expression of TNF-α and OX-42 over the dorsal spinal cord was substantially attenuated by hAFMSCs. The increased amplitude of sensory evoked potential as well as expression of synaptophysin and TNF-α expression was alleviated by hAFMSCs. Human AFMSCs significantly improved the threshold of mechanical allodynia and thermal hyperalgesia as well as various parameters of CatWalk XT gait analysis.Human AFMSCs administration could alleviate the neuropathic pain demonstrated in histomorphological alteration and neurobehavior possibly through the modulation of the inflammatory response.

  3. Combination of Local Transplantation of In Vitro Bone-marrow Stromal Cells and Pulsed Electromagnetic Fields Accelerate Functional Recovery of Transected Sciatic Nerve Regeneration: A Novel Approach in Transected Nerve Repair.

    Science.gov (United States)

    Mohammadi, Rahim; Mahmoodzadeh, Sirvan

    2015-01-01

    Effect of combination of undifferentiated bone marrow stromal cells (BMSCs) and pulsed electromagnetic fields (PEMF) on transected sciatic nerve regeneration was assessed in rats. A 10 mm nerve segment was excised and a vein graft was used to bridge the gap. Twenty microliter undifferentiated BMSCs (2× 107 cells /mL) were administered into the graft inBMSC group with no exposure to PEMF. In BMSC/PEMF group the whole body was exposed to PEMF (0.3 mT, 2Hz) for 4h/day within 1-5 days. In PEMF group the transected nerve was bridged and phosphate buffered saline was administered into the graft. In authograft group (AUTO), the transected nervesegments were reimplanted reversely and the whole body was exposed to PEMF. The regenerated nerve fibers were studied within 12 weeks after surgery. Behavioral, functional, electrophysiological, biomechanical, gastrocnemius muscle mass findings, morphometric indices and immuonohistochemical reactions confirmed faster recovery of regenerated axons in BMSC/PEMF group compared to those in the other groups (Pelectromagnetic fields could be considered as an effective, safe and tolerable treatment for peripheral nerve repair in clinical practice.

  4. Effect of alpha lipoic acid on retinal ganglion cell survival in an optic nerve crush model

    Science.gov (United States)

    Liu, Ruixing; Wang, Yanling; Pu, Mingliang

    2016-01-01

    Purpose This study was conducted to determine whether alpha lipoic acid (ALA) promotes the survival of retinal ganglion cells (RGCs) in a rat model of optic nerve crush (ONC) injury and to investigate the neuroprotective mechanisms of ALA in the retina in this ONC injury model. Methods Adult male Sprague-Dawley rats (180–220 g) were subjected to ONC injury surgery. ALA (63 mg/kg) was injected intravenously 1 day before or after the ONC injury. Animals were euthanized after 10 days, and the number of ganglion cells positive for RNA-binding protein with multiple splicing (Rbpms), which is an RGC marker, were counted on the whole mount retinas. In addition, immunofluorescence and immunoblotting were performed to examine the localization and levels of erythropoietin receptor (EPOR) and neurotrophin-4/5 (NT4/5) in the retinas in all experimental groups. To determine whether the EPO/EPOR signaling pathway was involved in the ALA antioxidant pathway, the rats were subjected to ruxolitinib (INCB018424, 0.25 mg/kg, bid, intraperitoneal, i.p.) treatment after the animals were injected intravenously with ALA 1 day before ONC injury. Results The average number of Rbpms-positive cells/mm2 in the control group (sham-operated group), the ONC group, the ALA-ONC group, and the ONC-ALA group retinas was 2219±28, 418±8, 848±22, and 613±18/mm2, respectively. The ALA-ONC and ONC-ALA groups showed a statistically significantly increased RGC survival rate compared to the ONC group. There were statistical differences in the RGC survival rates between the ALA-ONC (39%) and ONC-ALA groups (28%; p<0.05). Immunofluorescent labeling showed that EPOR and NT4/5 expression was significant in the retinal ganglion cell layer (GCL). At the same time, western blot analysis revealed that ALA induced upregulation of EPOR protein and NT4/5 protein expression in the retina after ONC injury. However, INCB018424 reversed the protective effects of ALA on the ONC retinas. Conclusions ALA has

  5. Human Beta Cells Produce and Release Serotonin to Inhibit Glucagon Secretion from Alpha Cells

    OpenAIRE

    Joana Almaça; Judith Molina; Danusa Menegaz; Pronin, Alexey N.; Alejandro Tamayo; Vladlen Slepak; Per-Olof Berggren; Alejandro Caicedo

    2016-01-01

    In the pancreatic islet, serotonin is an autocrine signal increasing beta cell mass during metabolic challenges such as those associated with pregnancy or high-fat diet. It is still unclear whether serotonin is relevant for regular islet physiology and hormone secretion. Here, we show that human beta cells produce and secrete serotonin when stimulated with increases in glucose concentration. Serotonin secretion from beta cells decreases cyclic AMP (cAMP) levels in neighboring alpha cells via ...

  6. Nerve injury induces glial cell line-derived neurotrophic factor (GDNF) expression in Schwann cells through purinergic signaling and the PKC-PKD pathway.

    Science.gov (United States)

    Xu, Pin; Rosen, Kenneth M; Hedstrom, Kristian; Rey, Osvaldo; Guha, Sushovan; Hart, Courtney; Corfas, Gabriel

    2013-07-01

    Upon peripheral nerve injury, specific molecular events, including increases in the expression of selected neurotrophic factors, are initiated to prepare the tissue for regeneration. However, the mechanisms underlying these events and the nature of the cells involved are poorly understood. We used the injury-induced upregulation of glial cell-derived neurotrophic factor (GDNF) expression as a tool to gain insights into these processes. We found that both myelinating and nonmyelinating Schwann cells are responsible for the dramatic increase in GDNF expression after injury. We also demonstrate that the GDNF upregulation is mediated by a signaling cascade involving activation of Schwann cell purinergic receptors, followed by protein kinase C signaling which activates protein kinase D (PKD), which leads to increased GDNF transcription. Given the potent effects of GDNF on survival and repair of injured peripheral neurons, we propose that targeting these pathways may yield therapeutic tools to treat peripheral nerve injury and neuropathies.

  7. Neural Stem Cell Affinity of Chitosan and Feasibility of Chitosan-Based Porous Conduits as Scaffolds for Nerve Tissue Engineering

    Institute of Scientific and Technical Information of China (English)

    WANG Aijun; AO Qiang; HE Qing; GONG Xiaoming; GONG Kai; GONG Yandao; ZHAO Nanming; ZHANG Xiufang

    2006-01-01

    Neural stem cells (NSCs) are currently considered as powerful candidate seeding cells for regeneration of both spinal cords and peripheral nerves. In this study, NSCs derived from fetal rat cortices were co-cultured with chitosan to evaluate the cell affinity of this material. The results showed that NSCs grew and proliferated well on chitosan films and most of them differentiated into neuron-like cells after 4 days of culture. Then, molded and braided chitosan conduits were fabricated and characterized for their cytotoxicity, swelling, and mechanical properties. Both types of conduits had no cytotoxic effects on fibroblasts (L929 cells) or neuroblastoma (Neuro-2a) cells. The molded conduits are much softer and more flexible while the braided conduits possess much better mechanical properties, which suggests different potential applications.

  8. [Trials of in vitro propagation of the scrapie agent in mouse nerve cells].

    Science.gov (United States)

    Markovits, P; Dormont, D; Delpech, B; Court, L; Latarjet, R

    1981-11-02

    We attempted to propagate the Scrapie agent in vitro in glia and neuroblastoma cells of Mice. Four out of seven assays of infection were positive, i.e. after several passages in vitro yielding at most a 10(9) fold final dilution of the original material, the extracts of each of the four cultures, when injected intracerebrally into CD1 Mice, produced a deadly disease displaying the clinical and pathological signs characteristic of Scrapie.

  9. To fabricate artificial nerves with tissue engineering methods

    Institute of Scientific and Technical Information of China (English)

    程飚; 陈峥嵘

    2002-01-01

    To fabricate artificial nerves with tissue engineering methods in vitro. Methods: Schwann cells (SCs) were cultured and seeded on polyglactin 910 fibers wrapped by biomembrane coated with rat tail glue and laminin for 2 weeks. The absorbability on the scaffolds, growth and migration of SCs were assessed with a light microscope, a scanning electron microscope and a transmission electron microscope. Results: SCs could migrate and proliferate on polyglactin 910 fibers. They were well distributed between scaffolds and absorbed on surface of scaffolds and formed a bungner band, on which SCs produced more matrices. SCs seeded on the biomembrane could also grow well. Axon regeneration in the distal nerve stump was observed at 8 weeks. Conclusions: Adult SCs can be expanded on coated fibers and biomembrane. Three-dimensional scaffold of SCs has the basic characteristics of artificial nerves. These findings offer a novel method to fabricate artificial nerves with tissue engineering methods for repairing defected long nerves.

  10. Increased encapsulated cell biodelivery of nerve growth factor in the brain by transposon-mediated gene transfer

    DEFF Research Database (Denmark)

    Fjord-Larsen, L; Kusk, Poul Henrik; Emerich, D F

    2012-01-01

    Nerve growth factor (NGF) is a potential therapeutic agent for Alzheimer's disease (AD) as it has positive effects on the basal forebrain cholinergic neurons whose degeneration correlates with the cognitive decline in AD. We have previously described an encapsulated cell biodelivery device, NsG0202......, capable of local delivery of NGF by a genetically modified human cell line, NGC-0295. The NsG0202 devices have shown promising safety and therapeutic results in a small phase 1b clinical study. However, results also show that the NGF dose could advantageously be increased. We have used the sleeping beauty...

  11. Release of chemical transmitters from cell bodies and dendrites of nerve cells

    OpenAIRE

    De-Miguel, Francisco F.; Nicholls, John G.

    2015-01-01

    Papers in this issue concern extrasynaptic transmission, namely release of signalling molecules by exocytosis or diffusion from neuronal cell bodies, dendrites, axons and glia. Problems discussed concern the molecules, their secretion and importance for normal function and disease. Molecules secreted extrasynaptically include transmitters, peptides, hormones and nitric oxide. For extrasynaptic secretion, trains of action potentials are required, and the time course of release is slower than a...

  12. Ultrastructural analysis of guided nerve regeneration using progesterone- and pregnenolone-loaded chitosan prostheses.

    Science.gov (United States)

    Chávez-Delgado, M E; Gomez-Pinedo, U; Feria-Velasco, A; Huerta-Viera, M; Castañeda, S Castro; Toral, F A López-Dellamary; Parducz, A; Anda, S Luquín-De; Mora-Galindo, J; García-Estrada, J

    2005-07-01

    Recently, numerous guide chambers for the treatment of injured nerves made up of different biomaterials have been designed, capable of hosting living cells or carrying neurotrophic or neuroactive substances to be directly released to the injured tissue. In this study, chitosan prostheses containing neurosteroids (progesterone and pregnenolone) were used for bridging a 10-mm gap in the rabbit facial nerve. Gas chromatography was used to quantify neurosteroid content in the prostheses prior to and after subcutaneous implantation at different periods of up to 60 days. The regeneration of the nerve fibers were evaluated at 15 and 45 days after axotomy by means of ultrastructural morphometric analysis. Different nerve fibers regenerative patterns were seen depending the groups studied and the analyzed stages. At 15 days after axotomy, the newly regenerating tissue revealed Schwann cells holding nonmyelinated nerve fiber bundles in an incipient and organized regenerative pattern. At 45 days, the regenerating tissue showed myelinated nerve fibers of different sizes, shapes, and myelin sheath thickness. Although the regeneration of the nerve fibers under neurosteroid treatment showed statistically significant differences in comparison with vehicle regenerated tissue, progesterone-loaded chitosan prostheses produced the best guided nerve regeneration response. These findings indicate that chitosan prostheses allowed regeneration of nerve fibers in their lumen, and when containing neurosteroids produced a faster guided nerve regeneration acting as a long-lasting release delivery vehicle.

  13. IL-10 and NOS2 modulate antigen-specific reactivity and nerve infiltration by T cells in experimental leprosy.

    Directory of Open Access Journals (Sweden)

    Deanna A Hagge

    2014-09-01

    Full Text Available Although immunopathology dictates clinical outcome in leprosy, the dynamics of early and chronic infection are poorly defined. In the tuberculoid region of the spectrum, Mycobacterium leprae growth is restricted yet a severe granulomatous lesion can occur. The evolution and maintenance of chronic inflammatory processes like those observed in the leprosy granuloma involve an ongoing network of communications via cytokines. IL-10 has immunosuppressive properties and IL-10 genetic variants have been associated with leprosy development and reactions.The role of IL-10 in resistance and inflammation in leprosy was investigated using Mycobacterium leprae infection of mice deficient in IL-10 (IL-10-/-, as well as mice deficient in both inducible nitric oxide synthase (NOS2-/- and IL-10 (10NOS2-/-. Although a lack of IL-10 did not affect M. leprae multiplication in the footpads (FP, inflammation increased from C57Bl/6 (B6producing CD4+CD44+ cells responding to M. leprae cell wall, membrane, and cytosol antigens and ML2028 (Ag85B were significantly increased in the evolved granuloma in NOS2-/- FP compared to B6 and IL-10-/- during early and peak phases. In 10NOS2-/- FP, CD4+CD44+ and especially CD8+CD44+ responses were augmented even further to these antigens as well as to ML0380 (GroES, ML2038 (bacterioferritin, and ML1877 (EF-Tu. Moreover, fragmented nerves containing CD4+ cells were present in 10NOS2-/- FP.The 10NOS2-/- strain offers insight on the regulation of granuloma formation and maintenance by immune modulators in the resistant forms of leprosy and presents a new model for investigating the pathogenesis of neurological involvement.

  14. Codonopsis pilosula (Franch) Nannftotal alkaloids potentiate neurite outgrowth induced by nerve growth factor in PC12 cells

    Institute of Scientific and Technical Information of China (English)

    LIUJian-Hui; BAOYong-Ming; SONGJi-Jun; ANLi-Jia

    2003-01-01

    AIM:To explore the effect of Codonopsis pilosula (Franch) Nannf total alkaloids (DSA) on differentiation inducedby nerve growth factor (NGF) in PC12 cells. METHODS: After culturing PC12 cells with DSA in the presence orabsence of NGF, neurite outgrowth in PC12 cells and correlated protein kinases were assayed. RESULTS: DSAalone did not exhibit neuritogenic activity, but caused a significant enhancement of NGF (2 μg/L)-induced neuriteoutgrowth in PC12 cells, and increased the phosphorylation of mitogen-activated protein kinase (MAPK).Furthermore, this enhancing effect was completely blocked by a specific MAPK kinase inhibitor, PD98059.CONCLUSION: DSA enhanced the NGF-induced neurite outgrowth in PC12 cells by amplifying an up-streamstep of the MAPK-dependent signaling pathway.

  15. Expression of nerve growth factor is upregulated in the rat thymic epithelial cells during thymus regeneration following acute thymic involution.

    Science.gov (United States)

    Lee, Hee-Woo; Kim, Sung-Min; Shim, Na-Ri; Bae, Soo-Kyung; Jung, Il-Gun; Kwak, Jong-Young; Kim, Bong-Seon; Kim, Jae-Bong; Moon, Jeon-Ok; Chung, Joo-Seop; Yoon, Sik

    2007-06-07

    Neuroimmune networks in the thymic microenvironment are thought to be involved in the regulation of T cell development. Nerve growth factor (NGF) is increasingly recognized as a potent immunomodulator, promoting "cross-talk" between various types of immune system cells. The present study describes the expression of NGF during thymus regeneration following acute involution induced by cyclophosphamide in the rat. Immunohistochemical stain demonstrated not only the presence of NGF but also its upregulated expression mainly in the subcapsular, paraseptal, and perivascular epithelial cells, and medullary epithelial cells including Hassall's corpuscles in both the normal and regenerating thymus. Biochemical data obtained using Western blot and RT-PCR supported these results and showed that thymic extracts contain NGF protein and mRNA, at higher levels during thymus regeneration. Thus, our results suggest that NGF expressed in these thymic epithelial cells plays a role in the T lymphopoiesis associated with thymus regeneration during recovery from acute thymic involution.

  16. Acellular nerve scaffold loading bone marrow mesenchymal stem cells to prepare artificial nerve for sciatic nerve defects%无细胞神经支架复合骨髓间充质干细胞构建组织工程人工神经修复坐骨神经缺损

    Institute of Scientific and Technical Information of China (English)

    张彩顺; 吕刚

    2011-01-01

    背景:作者已经成功制备了无细胞神经移植物,并且复合骨髓间充质干细胞构建组织工程人工神经桥接大鼠坐骨神经缺损.目的:无细胞神经移植物复合骨髓间充质干细胞构建组织工程人工神经修复大鼠坐骨神经缺损后运动功能的恢复.方法:成年雄性SD大鼠构建大鼠坐骨神经15 mm缺损模型,分别应用组织工程人工神经、组织工程神经支架或自行神经桥接坐骨神经缺损.桥接后20周再生神经电生理学测定,手术侧胫骨前肌湿质量、腓肠肌组织学及透视电镜分析.结果与结论:桥接20周后,组织工程人工神经与自体神经移植组胫骨前肌湿质量比较,差异无显著性意义(P > 0.05),神经干传导速度为(30.56±2.15)m/s.结果提示,无细胞神经移植物复合骨髓间充质干细胞构建的组织工程人工神经桥接大鼠坐骨神经缺损后,可以促进再生神经运动功能的恢复.%BACKGROUND: The authors have successfully prepared an acellular nerve graft to be combined with bone marrow mesenchymal stem cells in artificial nerve bridge of rat sciatic nerve defects.OBJECTIVE: To investigate the motor function of rats with sciatic nerve defects following repairing using acellular nerve graft combined with bone marrow mesenchymal stem cells.METHODS: Adult SD rats were used to prepare rat models of 15-mm sciatic nerve defects. Tissue-engineered nerve, tissue-engineered nerve scaffold, and autogenous nerve were separately applied to bridge sciatic nerve defects. After 20 weeks, electrophysiological determination of regenerated nerves was conducted, wet weight of the anterior tibial muscle was determined,and histological and transmission electron microscopy analyses of the gastrocnemius was done.RESULTS AND CONCLUSION: Twenty weeks after nerve grafting, no difference in the wet weight of the anterior tibial muscle was shown to be significant between tissue-engineered nerve and autogenous nerve grafts (P > 0

  17. Pro-nerve growth factor induces autocrine stimulation of breast cancer cell invasion through tropomyosin-related kinase A (TrkA) and sortilin protein.

    Science.gov (United States)

    Demont, Yohann; Corbet, Cyril; Page, Adeline; Ataman-Önal, Yasemin; Choquet-Kastylevsky, Genevieve; Fliniaux, Ingrid; Le Bourhis, Xuefen; Toillon, Robert-Alain; Bradshaw, Ralph A; Hondermarck, Hubert

    2012-01-13

    The precursor of nerve growth factor (proNGF) has been described as a biologically active polypeptide able to induce apoptosis in neuronal cells, via the neurotrophin receptor p75(NTR) and the sortilin receptor. Herein, it is shown that proNGF is produced and secreted by breast cancer cells, stimulating their invasion. Using Western blotting and mass spectrometry, proNGF was detected in a panel of breast cancer cells as well as in their conditioned media. Immunohistochemical analysis indicated an overproduction of proNGF in breast tumors, when compared with benign and normal breast biopsies, and a relationship to lymph node invasion in ductal carcinomas. Interestingly, siRNA against proNGF induced a decrease of breast cancer cell invasion that was restored by the addition of non-cleavable proNGF. The activation of TrkA, Akt, and Src, but not the MAP kinases, was observed. In addition, the proNGF invasive effect was inhibited by the Trk pharmacological inhibitor K252a, a kinase-dead TrkA, and siRNA against TrkA sortilin, neurotensin, whereas siRNA against p75(NTR) and the MAP kinase inhibitor PD98059 had no impact. These data reveal the existence of an autocrine loop stimulated by proNGF and mediated by TrkA and sortilin, with the activation of Akt and Src, for the stimulation of breast cancer cell invasion.

  18. Early transcutaneous electrical nerve stimulation reduces hyperalgesia and decreases activation of spinal glial cells in mice with neuropathic pain.

    Science.gov (United States)

    Matsuo, Hideaki; Uchida, Kenzo; Nakajima, Hideaki; Guerrero, Alexander Rodriguez; Watanabe, Shuji; Takeura, Naoto; Sugita, Daisuke; Shimada, Seiichiro; Nakatsuka, Terumasa; Baba, Hisatoshi

    2014-09-01

    Although transcutaneous electrical nerve stimulation (TENS) is widely used for the treatment of neuropathic pain, its effectiveness and mechanism of action in reducing neuropathic pain remain uncertain. We investigated the effects of early TENS (starting from the day after surgery) in mice with neuropathic pain, on hyperalgesia, glial cell activation, pain transmission neuron sensitization, expression of proinflammatory cytokines, and opioid receptors in the spinal dorsal horn. Following nerve injury, TENS and behavioral tests were performed every day. Immunohistochemical, immunoblot, and flow cytometric analysis of the lumbar spinal cord were performed after 8 days. Early TENS reduced mechanical and thermal hyperalgesia and decreased the activation of microglia and astrocytes (PEarly TENS decreased p-p38 within microglia (Pearly TENS relieved hyperalgesia in our mouse model of neuropathic pain by inhibiting glial activation, MAP kinase activation, PKC-γ, and p-CREB expression, and proinflammatory cytokines expression, as well as maintenance of spinal opioid receptors. The findings indicate that TENS treatment is more effective when applied as early after nerve injury as possible.

  19. Long-term administration of scopolamine interferes with nerve cell proliferation, differentiation and migration in adult mouse hippocampal dentate gyrus, but it does not induce cell death

    Institute of Scientific and Technical Information of China (English)

    Bing Chun Yan; Yun Lyul Lee; Il-Jun Kang; Moo-Ho Won; Joon Ha Park; Bai Hui Chen; Jeong-Hwi Cho; In Hye Kim; Ji Hyeon Ahn; Jae-Chul Lee; In Koo Hwang; Jun Hwi Cho

    2014-01-01

    Long-term administration of scopolamine, a muscarinic receptor antagonist, can inhibit the survival of newly generated cells, but its effect on the proliferation, differentiation and migration of nerve cells in the adult mouse hippocampal dentate gyrus remain poorly understood. In this study, we used immunohistochemistry and western blot methods to weekly detect the biological behaviors of nerve cells in the hippocampal dentate gyrus of adult mice that received intraperito-neal administration of scopolamine for 4 weeks. Expression of neuronal nuclear antigen (NeuN;a neuronal marker) and Fluoro-Jade B (a marker for the localization of neuronal degeneration) was also detected. After scopolamine treatment, mouse hippocampal neurons did not die, and Ki-67 (a marker for proliferating cells)-immunoreactive cells were reduced in number and reac hed the lowest level at 4 weeks. Doublecortin (DCX; a marker for newly generated neurons)-im-munoreactive cells were gradually shortened in length and reduced in number with time. After scopolamine treatment for 4 weeks, nearly all of the 5-bromo-2′-deoxyuridine (BrdU)-labeled newly generated cells were located in the subgranular zone of the dentate gyrus, but they did not migrate into the granule cell layer. Few mature BrdU/NeuN double-labeled cells were seen in the subgranular zone of the dentate gyrus. These ifndings suggest that long-term administration of scopolamine interferes with the proliferation, differentiation and migration of nerve cells in the adult mouse hippocampal dentate gyrus, but it does not induce cell death.

  20. 许旺细胞植入去细胞异体神经修复大鼠面神经缺损%Repair of facial nerve defects by using acellular nerve allografts implanted with Schwann cells in rats

    Institute of Scientific and Technical Information of China (English)

    朱国臣; 肖大江; 黄红宇; 袁渊; 吴四海; 赵新

    2008-01-01

    目的 观察体外分离、培养的许旺细胞对去细胞异体神经修复面神经长距离缺损的促进作用. 方法 30只大鼠随机分成A组、B组和c组(每组10只),制成而神经缺损12 mm模型,分别以去细胞异体神经联合许旺细胞、去细胞异体神经和自体神经移植修复.术后5个月分别行神经电生理、再生神经形态及数量等检测. 结果 A组神经肌肉动作电位的波幅恢复率(术侧/健侧)为(35.8±2.5)%,潜伏期恢复率(健侧/术侧)为(65.8±2.9)%;神经移植段再生轴突数为(1 570±188)个,髓鞘厚度为(0.383±0.031)μm.A组的各项指标优于B组(P0.05). 结论 许旺细胞植入去细胞异体神经可促进面神经长距离缺损的修复.%Objective To observe the effects of in vitro isolated Schwann cells co-cultured with chemically acellular nerve allografts on improving repair of large facial nerve defects. Methods A total of 30 Wistar rats were equally randomized into three groups, ie, experimental group, allograft group and autograft group. Nerve defect of 12 mm in length was made in the left inferior buccal branch of facial nerve and repaired with acellular nerve allograft implanted with Schwann cells, acellular nerve allograft and fresh tibial nerve autograft respectively. At the 5th month postoperatively, the function and morpholo-gy of the regenerated nerves were observed by electrophysiological method, methylene blue staining and transmission electron microscope. Results In experimental group, the recovery rate (operation side/normal side) of amplitude of nerve-muscle action potential was (35.8±2.5)%, the lantency recovery rate (normal side/operation side) (65.8±2.9)%, the number of the regenerated axon 1 570±188 and the myelin thickness (0.383±0.031) μm. The results in the experimental group were significantly supe-rior to those in the acellular nerve allograft group (P 0.05). Conclusion Transplantation of Schwarm cells in acellular nerve allograft can im

  1. Insulin-producing cells generated from dedifferentiated human pancreatic beta cells expanded in vitro.

    Directory of Open Access Journals (Sweden)

    Holger A Russ

    Full Text Available BACKGROUND: Expansion of beta cells from the limited number of adult human islet donors is an attractive prospect for increasing cell availability for cell therapy of diabetes. However, attempts at expanding human islet cells in tissue culture result in loss of beta-cell phenotype. Using a lineage-tracing approach we provided evidence for massive proliferation of beta-cell-derived (BCD cells within these cultures. Expansion involves dedifferentiation resembling epithelial-mesenchymal transition (EMT. Epigenetic analyses indicate that key beta-cell genes maintain open chromatin structure in expanded BCD cells, although they are not transcribed. Here we investigated whether BCD cells can be redifferentiated into beta-like cells. METHODOLOGY/PRINCIPAL FINDING: Redifferentiation conditions were screened by following activation of an insulin-DsRed2 reporter gene. Redifferentiated cells were characterized for gene expression, insulin content and secretion assays, and presence of secretory vesicles by electron microscopy. BCD cells were induced to redifferentiate by a combination of soluble factors. The redifferentiated cells expressed beta-cell genes, stored insulin in typical secretory vesicles, and released it in response to glucose. The redifferentiation process involved mesenchymal-epithelial transition, as judged by changes in gene expression. Moreover, inhibition of the EMT effector SLUG (SNAI2 using shRNA resulted in stimulation of redifferentiation. Lineage-traced cells also gave rise at a low rate to cells expressing other islet hormones, suggesting transition of BCD cells through an islet progenitor-like stage during redifferentiation. CONCLUSIONS/SIGNIFICANCE: These findings demonstrate for the first time that expanded dedifferentiated beta cells can be induced to redifferentiate in culture. The findings suggest that ex-vivo expansion of adult human islet cells is a promising approach for generation of insulin-producing cells for

  2. A Stem Cell-Seeded Nanofibrous Scaffold for Auditory Nerve Replacement

    Science.gov (United States)

    2013-10-01

    biopolymer scaffold within the internal auditory meatus (IAM) of the guinea pig. (A) The lateral wall of an intact guinea pig temporal bone is shown......Nanofibrous Scaffold for Auditory Nerve Replacement 5b. GRANT NUMBER W81XWH-12-1-0492 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S) Betty Diamond

  3. Nerve growth factor injected into the gastric ulcer base incorporates into endothelial, neuronal, glial and epithelial cells: implications for angiogenesis, mucosal regeneration and ulcer healing.

    Science.gov (United States)

    Tanigawa, T; Ahluwalia, A; Watanabe, T; Arakawa, T; Tarnawski, A S

    2015-08-01

    A previous study has demonstrated that locally administered growth factors such as epidermal growth factor, basic fibroblast growth factor and hepatocyte growth factor can accelerate healing of experimental gastric ulcers in rats. That study indicates that locally administered growth factors can exert potent biological effects resulting in enhanced gastric ulcers healing. However, the fate of injected growth factors, their retention and localization to specific cellular compartments have not been examined. In our preliminary study, we demonstrated that local injection of nerve growth factor to the base of experimental gastric ulcers dramatically accelerates ulcer healing, increases angiogenesis - new blood vessel formation, and improves the quality of vascular and epithelial regeneration. Before embarking on larger, definitive and time sequence studies, we wished to determine whether locally injected nerve growth factor is retained in gastric ulcer's tissues and taken up by specific cells during gastric ulcer healing. Gastric ulcers were induced in anesthetized rats by local application of acetic acid using standard methods; and, 60 min later fluorescein isothiocyanate-labeled nerve growth factor was injected locally to the ulcer base. Rats were euthanized 2, 5 and 10 days later. Gastric specimens were obtained and processed for histology. Unstained paraffin sections were examined under a fluorescence microscope, and the incorporation of fluorescein isothiocyanate-labeled nerve growth factor into various gastric tissue cells was determined and quantified. In addition, we performed immunostaining for S100β protein that is expressed in neural components. Five and ten days after ulcer induction labeled nerve growth factor (injected to the gastric ulcer base) was incorporated into endothelial cells of blood vessels, neuronal, glial and epithelial cells, myofibroblasts and muscle cells. This study demonstrates for the first time that during gastric ulcer healing

  4. Expression of NF-кB in Schwann cells and its effect on motor neuron apoptosis in spinal cord following sciatic nerves injury in rats

    Institute of Scientific and Technical Information of China (English)

    WANG Yong-tang; LU Xiu-min; YU Ying; YANG Yan-hong; GAO Jie

    2007-01-01

    Objective:To explore the expression of nuclear factor-kappa B (NF-кB) in Schwann cells (SCs) and its effect on motor neuron apoptosis in spinal cord following sciatic nerves injury in adult rats. Methods:Thirty-six adult Sprague-Dawley (SD) rats were divided randomly into normal control group (n=6),and sciatic nerves crushing group (n=30),and the later was further equally randomized into 5 nerves were examined by immunohistochemistry staining,and the apoptosis of motor neurons in spinalcord of lumbar 4 to lumbar 6(L4-L6)was investigated by terminal deoxynucleotidyl transferase mediated dUTP-biotin nick end labeling (TUNEL) assay.Both were quantitated by image analysis.Results:In the normal control group (P<0.05,P<0.01).At 1 d after sciatic nerves crushing,the expression of served in the time-course on motor neuron apoptosis after sciatic nerves injury.Correlation analyses refollowing sciatic nerves injury(r=0.976 0,P<0.01).Conclusion:After injury of sciatic nerves,the presence and up-regulation of NF-κB in SCs may be involved in motor neuron apoptosis in L4-L6 spinal cord.

  5. Mouse endometrial stromal cells produce basement-membrane components

    DEFF Research Database (Denmark)

    Wewer, U M; Damjanov, A; Weiss, J;

    1986-01-01

    During mouse pregnancy, uterine stromal cells transform into morphologically distinct decidual cells under the influence of the implanting embryo and a proper hormonal environment. Mechanical stimulation of hormonally primed uterine stromal cells leads to the same morphologic alterations. The dec...

  6. The role of cAMP in nerve growth factor-promoted neurite outgrowth in PC12 cells

    OpenAIRE

    1986-01-01

    Nerve growth factor (NGF)-mediated neurite outgrowth in rat pheochromocytoma PC12 cells has been described to be synergistically potentiated by the simultaneous addition of dibutyryl cAMP. To elucidate further the role of cAMP in NGF-induced neurite outgrowth we have used the adenylate cyclase activator forskolin, cAMP, and a set of chemically modified cAMP analogues, including the adenosine cyclic 3',5'-phosphorothioates (cAMPS) (Rp)-cAMPS and (Sp)-cAMPS. These diastereomers have differentia...

  7. Use of electrospinning to construct biomaterials for peripheral nerve regeneration.

    Science.gov (United States)

    Quan, Qi; Chang, Biao; Meng, Hao Ye; Liu, Ruo Xi; Wang, Yu; Lu, Shi Bi; Peng, Jiang; Zhao, Qing

    2016-10-01

    A number of limitations associated with the use of hollow nerve guidance conduits (NGCs) require further discussion. Most importantly, the functional recovery outcomes after the placement of hollow NGCs are poor even after the successful bridging of peripheral nerve injuries. However, nerve regeneration scaffolds built using electric spinning have several advantages that may improve functional recovery. Thus, the present study summarizes recent developments in this area, including the key cells that are combined with the scaffold and associated with nerve regeneration, the structure and configuration of the electrospinning design (which determines the performance of the electrospinning scaffold), the materials the electrospinning fibers are composed of, and the methods used to control the morphology of a single fiber. Additionally, this study also discusses the processes underlying peripheral nerve regeneration. The primary goals of the present review were to evaluate and consolidate the findings of studies that used scaffolding biomaterials built by electrospinning used for peripheral nerve regeneration support. It is amazing that the field of peripheral nerve regeneration continues to consistently produce such a wide variety of innovative techniques and novel types of equipment, because the introduction of every new process creates an opportunity for advances in materials for nerve repair.

  8. CAP1 was associated with actin and involved in Schwann cell differentiation and motility after sciatic nerve injury.

    Science.gov (United States)

    Zhu, Xinhui; Yao, Li; Guo, Aisong; Li, Aihong; Sun, Huiqing; Wang, Ning; Liu, Hanzhang; Duan, Zhiqin; Cao, Jianhua

    2014-06-01

    Adenylate cyclase-associated protein 1 (CAP1), a member of cyclase-associated proteins that regulating actin dynamics, was shown to regulate actin filaments, localize to dynamic actin structures and mediate such processes as establishment of cell polarity, motility, morphogenesis, receptor-mediated endocytosis and mRNA location. But little is known about the role of CAP1 during peripheral nervous system injury. Here, we found the spatiotemporal protein expression of CAP1 after sciatic nerve crush. After crush, CAP1 had an increased protein expression level, reached a peak at about day 5 and then returned to the normal level at 4 weeks, similar to Oct-6. Besides, in 5-day injured tissue, using double immunofluorescent staining we found CAP1 had a colocalization with S100 and Oct-6. In vitro, during the process of cAMP-induced Schwann cells differentiation, we observed enhanced expression of CAP1 and P0. Specially, CAP1-specific siRNA-tranfected SCs did not show significant actin structure which form cellure surface tension and protrusion shape after cAMP treatment. And we observed the interaction of CAP1 with actin and that CAP1-specific siRNA-transfected SCs had a decreased motility and migration. Together, all these data indicated that the change of CAP1 protein expression was associated with Schwann cells motility and differentiation after the crush of sciatic nerve.

  9. Combination of engineered neural cell adhesion molecules and GDF-5 for improved neurite extension in nerve guide concepts.

    Science.gov (United States)

    Niere, Marc; Braun, Bettina; Gass, Rea; Sturany, Sabine; Volkmer, Hansjürgen

    2006-06-01

    Current therapeutical approaches for the treatment of severe lesions in the peripheral nervous system rely on the use of autologous tissue or the body's own Schwann cells. However, these approaches are limited and alternative strategies for peripheral nerve regeneration are required. Here we evaluate combinations of a variety of neuronal regeneration factors including engineered cell adhesion molecules and growth factors in embryonic model neurons to test the possible improvement of artificial nerve guides by cooperative mechanisms. Cell adhesion molecules L1 and neurofascin synergistically promote neurite elongation. The outgrowth promoting properties of both proteins can be combined and further increased within one chimeric protein. Addition of growth and differentiation factor 5 (GDF-5) further enhances neurite outgrowth in a substrate-independent manner. This effect is not due to a protective mode of action of GDF-5 against pro-apoptotic stimuli. Consequently, the study supports the idea that different modes of action of pro-regenerative factors may contribute synergistically to neurite outgrowth and emphasizes the applicability of combinations of proteins specifically involved in development of the nervous system for therapeutical approaches.

  10. Use of hybrid chitosan membranes and human mesenchymal stem cells from the Wharton jelly of umbilical cord for promoting nerve regeneration in an axonotmesis rat model

    Institute of Scientific and Technical Information of China (English)

    Andrea G(a)rtner; Yuri Nakamura; Satoshi Hayakawa; Akiyoshi Osakah; Beatriz Porto; Ana Lúcia Luís; Artur SP Varej(a)o; Ana Colette Maurício; Tiago Pereira; Maria Jo(a)o Sim(o)es; Paulo AS Armada-da-Silva; Miguel L Fran(c)a; Rosa Sousa; Simone Bompasso; Stefania Raimondo; Yuki Shirosaki

    2012-01-01

    Many studies have been dedicated to the development of scaffolds for improving post-traumatic nerve regeneration.The goal of this study was to assess the effect on nerve regeneration,associating a hybrid chitosan membrane with non-differentiated human mesenchymal stem cells isolated from Wharton's jelly of umbilical cord,in peripheral nerve reconstruction after crush injury.Chromosome analysis on human mesenchymal stem cell line from Wharton's jelly was carried out and no structural alterations were found in metaphase.Chitosan membranes were previously tested in vitro,to assess their ability in supporting human mesenchymal stem cell survival,expansion,and differentiation.For the in vivo testing,Sasco Sprague adult rats were divided in 4 groups of 6 or 7 animals each:Group 1,sciatic axonotmesis injury without any other intervention (Group 1-Crush); Group 2,the axonotmesis lesion of 3 mm was infiltrated with a suspension of 1 250- 1 500 human mesenchymal stem cells (total volume of 50 μL) (Group 2-CrushCell); Group 3,axonotmesis lesion of 3 mm was enwrapped with a chitosan type Ⅲ membrane covered with a monolayer of non-differentiated human mesenchymal stem cells (Group 3-CrushChitlllCell) and Group 4,axonotmesis lesion of 3 mm was enwrapped with a chitosan type Ⅲ membrane (Group 4-CrushChitlll).Motor and sensory functional recovery was evaluated throughout a healing period of 12 weeks using sciatic functional index,static sciatic index,extensor postural thrust,and withdrawal reflex latency.Stereological analysis was carried out on regenerated nerve fibers.Results showed that infiltration of human mesenchymal stem cells,or the combination of chitosan membrane enwrapment and human mesenchymal stem cell enrichment after nerve crush injury provide a slight advantage to post-traumatic nerve regeneration.Results obtained with chitosan type Ⅲ membrane alone confirmed that they significantly improve post-traumatic axonal regrowth and may represent a very promising

  11. Rgcs1, a dominant QTL that affects retinal ganglion cell death after optic nerve crush in mice

    Directory of Open Access Journals (Sweden)

    Schlamp Cassandra L

    2008-07-01

    Full Text Available Abstract Background Intrinsic apoptosis of neuronal somas is one aspect of neurodegenerative diseases that can be influenced by genetic background. Genes that affect this process may act as susceptibility alleles that contribute to the complex genetic nature of these diseases. Retinal ganglion cell death is a defining feature of the chronic and genetically complex neurodegenerative disease glaucoma. Previous studies using an optic nerve crush procedure in inbred mice, showed that ganglion cell resistance to crush was affected by the Mendelian-dominant inheritance of 1–2 predicted loci. To assess this further, we bred and phenotyped a large population of F2 mice derived from a resistant inbred strain (DBA/2J and a susceptible strain (BALB/cByJ. Results Genome wide mapping of the F2 mice using microsatellite markers, detected a single highly significant quantitative trait locus in a 25 cM (58 Mb interval on chromosome 5 (Chr5.loc34-59 cM. No interacting loci were detected at the resolution of this screen. We have designated this locus as Retinal ganglion cell susceptible 1, Rgcs1. In silico analysis of this region revealed the presence of 578 genes or expressed sequence tags, 4 of which are highly expressed in the ganglion cell layer of the mammalian retina, and 2 of which are suspected susceptibility alleles in chronic neurodegenerative diseases. In addition, 25 genes contain 36 known single nucleotide polymorphisms that create nonsynonymous amino acid changes between the two parental strains. Collectively, this analysis has identified 7 potential candidate genes that may affect ganglion cell death. Conclusion The process of ganglion cell death is likely one of the many facets of glaucoma susceptibility. A novel dominant locus has been identified that affects sensitivity of ganglion cells to optic nerve crush. The allele responsible for this sensitivity may also be a susceptibility allele for glaucoma.

  12. Efficient Differentiation of Mouse Embryonic Stem Cells into Insulin-Producing Cells

    Directory of Open Access Journals (Sweden)

    Szu-Hsiu Liu

    2012-01-01

    Full Text Available Embryonic stem (ES cells are a potential source of a variety of differentiated cells for cell therapy, drug discovery, and toxicology screening. Here, we present an efficacy strategy for the differentiation of mouse ES cells into insulin-producing cells (IPCs by a two-step differentiation protocol comprising of (i the formation of definitive endoderm in monolayer culture by activin A, and (ii this monolayer endoderm being induced to differentiate into IPCs by nicotinamide, insulin, and laminin. Differentiated cells can be obtained within approximately 7 days. The differentiation IPCs combined application of RT-PCR, ELISA, and immunofluorescence to characterize phenotypic and functional properties. In our study, we demonstrated that IPCs produced pancreatic transcription factors, endocrine progenitor marker, definitive endoderm, pancreatic β-cell markers, and Langerhans α and δ cells. The IPCs released insulin in a manner that was dose dependent upon the amount of glucose added. These techniques may be able to be applied to human ES cells, which would have very important ramifications for treating human disease.

  13. 骨髓间充质干细胞向神经细胞的分化%Bone marrow mesenchymal stem cells differentiation into nerve cells

    Institute of Scientific and Technical Information of China (English)

    安秀峰; 黄汉昌; 姜招峰

    2013-01-01

    近年来,骨髓间充质干细胞逐渐成为神经科学领域的研究热点,广泛用于治疗神经退行性疾病,其原因是;骨髓间充质干细胞可在诱导物存在下定向分化为有功能的神经元细胞,并能成功表达神经标志蛋白.结合近几年的研究进展,主要从骨髓间充质干细胞的临床试验和应用;激光辐射、氧气含量和神经细胞对骨髓间充质干细胞分化为神经细胞的影响;microRNA、Notch信号通路和Wnt信号通路对骨髓间充质干细胞分化为神经细胞的调节作用等三方面进行阐述.%For the past few years, bone marrow mesenchymal stem cells have gradually become a research focus in the field of neuroscience, and have been widely used in the treatment of neurodegenerative diseases. It is just because that it can differentiate into functional neuron and express neural marker proteins successfully when inducer exists. Combined with the recent research progress, this paper will introduce the bone marrow mesenchymal stem cells from the following three aspects: firstly, the clinical trial and application of bone marrow mesenchymal stem cells; secondly, the effects of laser radiation, oxygen content and the nerve cells on the process of bone marrow mesenchymal stem cells differentiate into nerve cells; thirdly, microRNA, Notch signal pathway and Wnt signal pathway regulate the process of bone marrow mesenchymal stem cells differentiating into nerve cells.

  14. Reversal of diabetes with insulin-producing cells derived in vitro from human pluripotent stem cells.

    Science.gov (United States)

    Rezania, Alireza; Bruin, Jennifer E; Arora, Payal; Rubin, Allison; Batushansky, Irina; Asadi, Ali; O'Dwyer, Shannon; Quiskamp, Nina; Mojibian, Majid; Albrecht, Tobias; Yang, Yu Hsuan Carol; Johnson, James D; Kieffer, Timothy J

    2014-11-01

    Transplantation of pancreatic progenitors or insulin-secreting cells derived from human embryonic stem cells (hESCs) has been proposed as a therapy for diabetes. We describe a seven-stage protocol that efficiently converts hESCs into insulin-producing cells. Stage (S) 7 cells expressed key markers of mature pancreatic beta cells, including MAFA, and displayed glucose-stimulated insulin secretion similar to that of human islets during static incubations in vitro. Additional characterization using single-cell imaging and dynamic glucose stimulation assays revealed similarities but also notable differences between S7 insulin-secreting cells and primary human beta cells. Nevertheless, S7 cells rapidly reversed diabetes in mice within 40 days, roughly four times faster than pancreatic progenitors. Therefore, although S7 cells are not fully equivalent to mature beta cells, their capacity for glucose-responsive insulin secretion and rapid reversal of diabetes in vivo makes them a promising alternative to pancreatic progenitor cells or cadaveric islets for the treatment of diabetes.

  15. Fibroblastic reticular cells from lymph nodes attenuate T cell expansion by producing nitric oxide.

    Directory of Open Access Journals (Sweden)

    Stefanie Siegert

    Full Text Available Adaptive immune responses are initiated when T cells encounter antigen on dendritic cells (DC in T zones of secondary lymphoid organs. T zones contain a 3-dimensional scaffold of fibroblastic reticular cells (FRC but currently it is unclear how FRC influence T cell activation. Here we report that FRC lines and ex vivo FRC inhibit T cell proliferation but not differentiation. FRC share this feature with fibroblasts from non-lymphoid tissues as well as mesenchymal stromal cells. We identified FRC as strong source of nitric oxide (NO thereby directly dampening T cell expansion as well as reducing the T cell priming capacity of DC. The expression of inducible nitric oxide synthase (iNOS was up-regulated in a subset of FRC by both DC-signals as well as interferon-γ produced by primed CD8+ T cells. Importantly, iNOS expression was induced during viral infection in vivo in both LN FRC and DC. As a consequence, the primary T cell response was found to be exaggerated in Inos(-/- mice. Our findings highlight that in addition to their established positive roles in T cell responses FRC and DC cooperate in a negative feedback loop to attenuate T cell expansion during acute inflammation.

  16. The effects of functional magnetic nanotubes with incorporated nerve growth factor in neuronal differentiation of PC12 cells

    Energy Technology Data Exchange (ETDEWEB)

    Xie Jining; Chen Linfeng; Varadan, Vijay K [Nanomaterials and Nanotubes Research Laboratory, College of Engineering, University of Arkansas, Fayetteville, AR 72701 (United States); Yancey, Justin; Srivatsan, Malathi [Department of Biological Sciences, Arkansas State University, State University, AR 72467 (United States)], E-mail: jxie@uark.edu, E-mail: msrivatsan@astate.edu

    2008-03-12

    In this in vitro study the efficiency of magnetic nanotubes to bind with nerve growth factor (NGF) and the ability of NGF-incorporated magnetic nanotubes to release the bound NGF are investigated using rat pheochromocytoma cells (PC12 cells). It is found that functional magnetic nanotubes with NGF incorporation enabled the differentiation of PC12 cells into neurons exhibiting growth cones and neurite outgrowth. Microscope observations show that filopodia extending from neuron growth cones were in close proximity to the NGF-incorporated magnetic nanotubes, at times appearing to extend towards or into them. These results show that magnetic nanotubes can be used as a delivery vehicle for NGF and thus may be exploited in attempts to treat neurodegenerative disorders such as Parkinson's disease with neurotrophins. Further neurite outgrowth can be controlled by manipulating magnetic nanotubes with external magnetic fields, thus helping in directed regeneration.

  17. Induction of autoantibody-producing cells after the coculture of haptenated and normal human mononuclear leukocytes.

    Science.gov (United States)

    Pisko, E J; Foster, S L; Turner, R A

    1981-10-01

    The coculture of normal human peripheral blood mononuclear leukocytes (PBL) and autologous mononuclear leukocytes coupled to the trinitrophenyl (TNP) hapten (TNP-PBL) was found to induce a polyclonal activation of antibody-producing cells. The polyclonal activation of antibody-producing cells was demonstrated by detecting the induction of cells producing antibody to sheep red blood cells using a complement-dependent, direct, hemolytic plaque-forming cell (PFC) assay. A ratio of four normal to one haptenated mononuclear leukocyte was found to be optimal for inducing the polyclonal activation of antibody-producing cell in these cultures. The plaque-forming cells assay in these experiments utilized monolayers of indicator red cells. Further evidence for the polyclonal induction of antibody-producing cells by TNP-PBL was provided by demonstrating PFC on monolayers of not only sheep red blood cells, but also autologous human red cells, bromelain-treated autologous red cells, TNP-coupled human and sheep red cells, and human autologous red cells coupled to human heat-aggregated IgG with chromic chloride. Thus cells secreting antibody to TNP, human red cells, and human IgG were induced. Anti-IgG and anti-human red cell-producing cells were first detected on Day 2 of culture and were still present on Day 9. Mononuclear leukocytes altered by chemical haptenation polyclonally stimulate normal mononuclear leukocytes to become antibody-producing cells. This polyclonal stimulation of antibody-producing cells includes cells producing antibodies to human IgG and human autologous red blood cells suggesting that autoantibody-producing cells are induced.

  18. Hypoxia-Induced Upregulation of miR-132 Promotes Schwann Cell Migration After Sciatic Nerve Injury by Targeting PRKAG3.

    Science.gov (United States)

    Yao, Chun; Shi, Xiangxiang; Zhang, Zhanhu; Zhou, Songlin; Qian, Tianmei; Wang, Yaxian; Ding, Fei; Gu, Xiaosong; Yu, Bin

    2016-10-01

    Following peripheral nerve injury, hypoxia is formed as a result of defects in blood supply at the injury site. Despite accumulating evidence on the effects of microRNAs (miRNAs) on phenotype modulation of Schwann cells (SCs) after peripheral nerve injury, the impact of hypoxia on SC behaviors through miRNAs during peripheral nerve regeneration has not been estimated. In this study, we confirmed our previous microarray data on the upregulation of miR-132 after sciatic nerve injury in rats and observed that overexpression of miR-132 significantly promoted cell migration of primary cultured SCs. Interestingly, hypoxia-increased expression of miR-132 also enhanced SC migration while inhibition of miR-132 suppressed hypoxia-induced increase in SC migration. miR-132 downregulated PRKAG3 through binding to its 3'-UTR, and PRKAG3 knockdown compromised the reducing effect of miR-132 inhibition on SC migration under normal or hypoxia condition. Moreover, in vivo injection of miR-132 agomir into rats with sciatic nerve transection accelerated SC migration from the proximal to distal stump. Overall, our results suggest that the hypoxia-induced upregulation of miR-132 could promote SC migration and facilitate peripheral nerve regeneration.

  19. Human Liver Cells Expressing Albumin and Mesenchymal Characteristics Give Rise to Insulin-Producing Cells

    Directory of Open Access Journals (Sweden)

    Irit Meivar-Levy

    2011-01-01

    Full Text Available Activation of the pancreatic lineage in the liver has been suggested as a potential autologous cell replacement therapy for diabetic patients. Transcription factors-induced liver-to-pancreas reprogramming has been demonstrated in numerous species both in vivo and in vitro. However, human-derived liver cells capable of acquiring the alternate pancreatic repertoire have never been characterized. It is yet unknown whether hepatic-like stem cells or rather adult liver cells give rise to insulin-producing cells. Using an in vitro experimental system, we demonstrate that proliferating adherent human liver cells acquire mesenchymal-like characteristics and a considerable level of cellular plasticity. However, using a lineage-tracing approach, we demonstrate that insulin-producing cells are primarily generated in cells enriched for adult hepatic markers that coexpress both albumin and mesenchymal markers. Taken together, our data suggest that adult human hepatic tissue retains a substantial level of developmental plasticity, which could be exploited in regenerative medicine approaches.

  20. Generation of insulin-producing cells from gnotobiotic porcine skin-derived stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Ji Hoon; Lee, Sung Ho; Heo, Young Tae [Department of Bioscience and Biotechnology, Bio-Organ Research Center, Konkuk University, Seoul 143-701 (Korea, Republic of); Uhm, Sang Jun [Department of Animal Biotechnology, Bio-Organ Research Center, Konkuk University, Seoul 143-701 (Korea, Republic of); Lee, Hoon Taek, E-mail: htl3675@konkuk.ac.kr [Department of Animal Biotechnology, Bio-Organ Research Center, Konkuk University, Seoul 143-701 (Korea, Republic of)

    2010-07-09

    A major problem in the treatment of type 1 diabetes mellitus is the limited availability of alternative sources of insulin-producing cells for islet transplantation. In this study, we investigated the effect of bone morphogenetic protein 4 (BMP-4) treatments of gnotobiotic porcine skin-derived stem cells (gSDSCs) on their reprogramming and subsequent differentiation into insulin-producing cells (IPCs). We isolated SDSCs from the ear skin of a gnotobiotic pig. During the proliferation period, the cells expressed stem-cell markers Oct-4, Sox-2, and CD90; nestin expression also increased significantly. The cells could differentiate into IPCs after treatments with activin-A, glucagon-like peptide-1 (GLP-1), and nicotinamide. After 15 days in the differentiation medium, controlled gSDSCs began expressing endocrine progenitor genes and proteins (Ngn3, Neuro-D, PDX-1, NKX2.2, NKX6.1, and insulin). The IPCs showed increased insulin synthesis after glucose stimulation. The results indicate that stem cells derived from the skin of gnotobiotic pigs can differentiate into IPCs under the appropriate conditions in vitro. Our three-stage induction protocol could be applied without genetic modification to source IPCs from stem cells in the skin of patients with diabetes for autologous transplantation.

  1. Highly efficient differentiation of human ES cells and iPS cells into mature pancreatic insulin-producing cells

    Institute of Scientific and Technical Information of China (English)

    Donghui Zhang; Wei Jiang; Meng Liu; Xin Sui; Xiaolei Yin; Song Chen; Yan Shi; Hongkui Deng

    2009-01-01

    Human pluripotent stem cells represent a potentially unlimited source of functional pancreatic endocrine lineage cells. Here we report a highly efficient approach to induce human embryonic stem (ES) cells and induced pluripotent stem (iPS) cells to differentiate into mature insulin-producing cells in a chemical-defined culture system. The differentiated human ES cells obtained by this approach comprised nearly 25% insulin-positive cells as assayed by flow cytometry analysis, which released insulin/C-peptide in response to glucose stimuli in a manner comparable to that of adult human islets. Most of these insulin-producing cells co-expressed mature β cell-specific markers such as NKX6-1 and PDX1, indicating a similar gene expression pattern to adult islet β cells in vivo. In this study, we also demonstrated that EGF facilitates the expansion of PDX1-positive pancreatic progenitors. Moreover, our protocol also succeeded in efficiently inducing human iPS cells to differentiate into insulin-producing cells. Therefore, this work not only provides a new model to study the mechanism of human pancreatic specialization and maturation in vitro, but also enhances the possibility of utilizing patient-specific iPS cells for the treatment of diabetes.

  2. Age-Related Yield of Adipose-Derived Stem Cells Bearing the Low-Affinity Nerve Growth Factor Receptor

    Directory of Open Access Journals (Sweden)

    Raquel Cuevas-Diaz Duran

    2013-01-01

    Full Text Available Adipose-derived stem cells (ADSCs are a heterogeneous cell population that may be enriched by positive selection with antibodies against the low-affinity nerve growth factor receptor (LNGFR or CD271, yielding a selective cell universe with higher proliferation and differentiation potential. This paper addresses the need for determining the quantity of ADSCs positive for the CD271 receptor and its correlation with donor's age. Mononuclear cells were harvested from the lower backs of 35 female donors and purified using magnetic beads. Multipotency capacity was tested by the expression of stemness genes and through differentiation into preosteoblasts and adipocytes. A significant statistical difference was found in CD271+ concentrations between defined age intervals. The highest yield was found within women on the 30–40-year-old age range. CD271+ ADSCs from all age groups showed differentiation capabilities as well as expression of typical multipotent stem cell genes. Our data suggest that the amount of CD271+ cells correlates inversely with age. However, the ability to obtain these cells was maintained through all age ranges with a yield higher than what has been reported from bone marrow. Our findings propose CD271+ ADSCs as the primary choice for tissue regeneration and autologous stem cell therapies in older subjects.

  3. Impulse noise transiently increased the permeability of nerve and glial cell membranes, an effect accentuated by a recent brain injury.

    Science.gov (United States)

    Säljö, Annette; Huang, Ying-Lai; Hansson, Hans-Arne

    2003-08-01

    A single exposure to intense impulse noise may cause diffuse brain injury, revealed by increased expression of immediate early gene products, transiently altered distribution of neurofilaments, accumulation of beta-amyloid precursor protein, apoptosis, and gliosis. Neither hemorrage nor any gross structural damage are seen. The present study focused on whether impulse noise exposure increased the permeability of nerve and glial cell membranes to proteins. Also, we investigated whether a preceding, minor focal surgical brain lesion accentuated the leakage of cytosolic proteins. Anaesthetized rats were exposed to a single impulse noise at either 199 or 202 dB for 2 milliseconds. Transiently elevated levels of the cellular protein neuron specific enolase (NSE) and the glial cytoplasmic protein S-100 were recorded in the cerebrospinal fluid (CSF) during the first hours after the exposure to 202 dB. A surgical brain injury, induced the day before the exposure to the impulse noise, was associated with significantly increased concentrations of both markers in the CSF. It is concluded that intense impulse noise damages both nerve and glial cells, an effect aggravated by a preexisting surgical lesion. The impulse of the shock wave, i.e. the pressure integrated over time, is likely to be the injurious mechanism. The abnormal membrane permeability and the associated cytoskeletal changes may initiate events, which eventually result in a progressive diffuse brain injury.

  4. Osteogenic Cells Derived From Embryonic Stem Cells Produced Bone Nodules in Three-Dimensional Scaffolds

    Directory of Open Access Journals (Sweden)

    Chaudhry G. R.

    2004-01-01

    Full Text Available An approach for 3D bone tissue generation from embryonic stem (ES cells was investigated. The ES cells were induced to differentiate into osteogenic precursors, capable of proliferating and subsequently differentiating into bone-forming cells. The differentiated cells and the seeded scaffolds were characterized using von Kossa and Alizarin Red staining, electron microscopy, and RT-PCR analysis. The results demonstrated that ES-derived bone-forming cells attached to and colonized the biocompatible and biodegradable scaffolds. Furthermore, these cells produced bone nodules when grown for 3–4 weeks in mineralization medium containing ascorbic acid and beta-glycerophosphate both in tissue culture plates and in scaffolds. The differentiated cells also expressed osteospecific markers when grown both in the culture plates and in 3D scaffolds. Osteogenic cells expressed alkaline phosphatase, osteocalcin, and osteopontin, but not an ES cell-specific marker, oct-4. These findings suggest that ES cell can be used for in vitro tissue engineering and cultivation of graftable skeletal structures.

  5. Imbalance between IL-17A-Producing Cells and Regulatory T Cells during Ischemic Stroke

    Directory of Open Access Journals (Sweden)

    Yuehua Hu

    2014-01-01

    Full Text Available Immune responses and inflammation are key elements in the pathogenesis of ischemic stroke (IS. Although the involvement of IL-17A in IS has been demonstrated using animal models, the involvement of IL-17A and IL-17-secreting T cell subsets in IS patients has not been verified, and whether the balance of Treg/IL-17-secreting T cells is altered in IS patients remains unknown. In the present study, we demonstrated that the proportion of peripheral Tregs and the levels of IL-10 and TGF-β were reduced in patients with IS compared with controls using flow cytometry (FCM, real-time PCR, and ELISA assays. However, the proportions of Th17 and γδ T cells, the primary IL-17A-secreting cells, increased dramatically, and these effects were accompanied by increases in the levels of IL-17A, IL-23, IL-6, and IL-1β in IS patients. These studies suggest that the increase in IL-17A-producing cells and decrease in Treg cells might contribute to the pathogenesis of IS. Manipulating the balance between Tregs and IL-17A-producing cells might be helpful for the treatment of IS.

  6. Activated Schwann Cell-Like Cells on Aligned Fibrin-Poly(Lactic-Co-Glycolic Acid) Structures: A Novel Construct for Application in Peripheral Nerve Regeneration.

    Science.gov (United States)

    Schuh, Christina M A P; Morton, Tatjana J; Banerjee, Asmita; Grasl, Christian; Schima, Heinrich; Schmidhammer, Robert; Redl, Heinz; Ruenzler, Dominik

    2015-01-01

    Tissue engineering approaches in nerve regeneration search for ways to support gold standard therapy (autologous nerve grafts) and to improve results by bridging nerve defects with different kinds of conduits. In this study, we describe electrospinning of aligned fibrin-poly(lactic-co-glycolic acid) (PLGA) fibers in an attempt to create a biomimicking tissue-like material seeded with Schwann cell-like cells (SCLs) in vitro for potential use as an in vivo scaffold. Rat adipose-derived stem cells (rASCs) were differentiated into SCLs and evaluated with flow cytometry concerning their differentiation and activation status [S100b, P75, myelin-associated glycoprotein (MAG), and protein 0 (P0)]. After receiving the proliferation stimulus forskolin, SCLs expressed S100b and P75; comparable to native, activated Schwann cells, while cultured without forskolin, cells switched to a promyelinating phenotype and expressed S100b, MAG, and P0. Human fibrinogen and thrombin, blended with PLGA, were electrospun and the alignment and homogeneity of the fibers were proven by scanning electron microscopy. Electrospun scaffolds were seeded with SCLs and the formation of Büngner-like structures in SCLs was evaluated with phalloidin/propidium iodide staining. Carrier fibrin gels containing rASCs acted as a self-shaping matrix to form a tubular structure. In this study, we could show that rASCs can be differentiated into activated, proliferating SCLs and that these cells react to minimal changes in stimulus, switching to a promyelinating phenotype. Aligned electrospun fibrin-PLGA fibers promoted the formation of Büngner-like structures in SCLs, which also rolled the fibrin-PLGA matrix into a tubular scaffold. These in vitro findings favor further in vivo testing.

  7. Progesterone and peripheral nerve regeneration

    Institute of Scientific and Technical Information of China (English)

    Fei Fan; Haichao Li; Yuwei Wang; Yanglin Zheng; Lianjun Jia; Zhihui Wang

    2006-01-01

    OBJECTIVE: To explore the effect of progesterone on peripheral nerve regeneration.DATA SOURCES: An online search of Medline and OVID databases was under taken to identify articles about progesterone and peripheral nerve regeneration published in English between January 1990 and June 2004 by using the keywords of "peripheral nerve, injury, progesterone, regeneration".STUDY SELECTION: The data were primarily screened, those correlated with progesterone and peripheral nerve regeneration were involved, and their original articles were further searched, the repetitive studies or reviews were excluded.DATA EXTRACTION: Totally 59 articles about progesterone and peripheral nerve regeneration were collected, and 26 of them were involved, the other 33 excluded ones were the repetitive studies or reviews.DATA SYNTHESIS: Recent researches found that certain amount of progesterone could be synthetized in peripheral nervous system, and the expression of progesterone receptor could be found in sensory neurons and Schwann cells. After combined with the receptor, endogenous and exogenous progesterone can accelerate the formation of peripheral nerve myelin sheath, also promote the axonal regeneration.CONCLUSION: Progesterone plays a role in protecting neurons, increasing the sensitivity of nerve tissue to nerve growth factor, and accelerating regeneration of nerve in peripheral nerve regeneration, which provides theoretical references for the treatment of demyelinated disease and nerve injury, as well as the prevention of neuroma, especially that the in vivo level of progesterone should be considered for the elderly people accompanied by neuropathy and patients with congenital luteal phase defect, which is of positive significance in guiding the treatment.

  8. Effects of umbilical cord tissue mesenchymal stem cells (UCX® on rat sciatic nerve regeneration after neurotmesis injuries

    Directory of Open Access Journals (Sweden)

    Gärtner A

    2013-04-01

    Full Text Available Peripheral nerves have the intrinsic capacity of self-regeneration after traumatic injury but the extent of the regeneration is often very poor. Increasing evidence demonstrates that mesenchymal stem/stromal cells (MSCs may play an important role in tissue regeneration through the secretion of soluble trophic factors that enhance and assist in repair by paracrine activation of surrounding cells. In the present study, the therapeutic value of a population of umbilical cord tissue-derived MSCs, obtained by a proprietary method (UCX®, was evaluated on end-to-end rat sciatic nerve repair. Furthermore, in order to promote both, end-to-end nerve fiber contacts and MSC cell-cell interaction, as well as reduce the flush away effect of the cells after administration, a commercially available haemostatic sealant, Floseal®, was used as vehicle. Both, functional and morphologic recoveries were evaluated along the healing period using extensor postural thrust (EPT, withdrawal reflex latency (WRL, ankle kinematics analysis, and either histological analysis or stereology, in the hyper-acute, acute and chronic phases of healing. The histological analysis of the hyper-acute and acute phase studies revealed that in the group treated with UCX ® alone the Wallerian degeneration was improved for the subsequent process of regeneration, the fiber organization was higher, and the extent of fibrosis was lower. The chronic phase experimental groups revealed that treatment with UCX® induced an increased number of regenerated fibers and thickening of the myelin sheet. Kinematics analysis showed that the ankle joint angle determined for untreated animals was significantly different from any of the treated groups at the instant of initial contact (IC. At opposite toe off (OT and heel rise (HR, differences were found between untreated animals and the groups treated with either UCX® alone or UCX® administered with Floseal®. Overall, the UCX® application presented

  9. Loss of Ab-nerve endings associated with the Merkel cell-neurite complex in the lesional oral mucosa epithelium of lichen planus and hyperkeratosis

    Institute of Scientific and Technical Information of China (English)

    Daniela Caldero n Carrio n; Yu ksel Korkmaz; Britta Cho; Marion Kopp; Wilhelm Bloch; Klaus Addicks; Wilhelm Niedermeier

    2016-01-01

    The Merkel cell-neurite complex initiates the perception of touch and mediates Ab slowly adapting type I responses. Lichen planus is a chronic inflammatory autoimmune disease with T-cell-mediated inflammation, whereas hyperkeratosis is characterized with or without epithelial dysplasia in the oral mucosa. To determine the effects of lichen planus and hyperkeratosis on the Merkel cell-neurite complex, healthy oral mucosal epithelium and lesional oral mucosal epithelium of lichen planus and hyperkeratosis patients were stained by immunohistochemistry (the avidin-biotin-peroxidase complex and double immunofluorescence methods) using pan cytokeratin, 20 (K20, a Merkel cell marker), and neurofilament 200 (NF200, a myelinated Ab- and Ad-nerve fibre marker) antibodies. NF200-immunoreactive (ir) nerve fibres in healthy tissues and in the lesional oralmucosa epitheliumof lichen planus and hyperkeratosis were counted and statistically analysed. In the healthy oral mucosa, K20-positive Merkel cells with and without close association to the intraepithelial NF200-ir nerve fibres were detected. In the lesional oral mucosa of lichen planus and hyperkeratosis patients, extremely rare NF200-ir nerve fibres were detected only in the lamina propria. Compared with healthy tissues, lichen planus and hyperkeratosis tissues had significantly decreased numbers of NF200-ir nerve fibres in the oral mucosal epithelium. Lichen planus and hyperkeratosis were associated with the absence of Ab-nerve endings in the oral mucosal epithelium. Thus, we conclude that mechanosensation mediated by the Merkel cell-neurite complex in the oral mucosal epithelium is impaired in lichen planus and hyperkeratosis.

  10. Loss of Aβ-nerve endings associated with the Merkel cell-neurite complex in the lesional oral mucosa epithelium of lichen planus and hyperkeratosis.

    Science.gov (United States)

    Carrión, Daniela Calderón; Korkmaz, Yüksel; Cho, Britta; Kopp, Marion; Bloch, Wilhelm; Addicks, Klaus; Niedermeier, Wilhelm

    2016-03-30

    The Merkel cell-neurite complex initiates the perception of touch and mediates Aβ slowly adapting type I responses. Lichen planus is a chronic inflammatory autoimmune disease with T-cell-mediated inflammation, whereas hyperkeratosis is characterized with or without epithelial dysplasia in the oral mucosa. To determine the effects of lichen planus and hyperkeratosis on the Merkel cell-neurite complex, healthy oral mucosal epithelium and lesional oral mucosal epithelium of lichen planus and hyperkeratosis patients were stained by immunohistochemistry (the avidin-biotin-peroxidase complex and double immunofluorescence methods) using pan cytokeratin, cytokeratin 20 (K20, a Merkel cell marker), and neurofilament 200 (NF200, a myelinated Aβ- and Aδ-nerve fibre marker) antibodies. NF200-immunoreactive (ir) nerve fibres in healthy tissues and in the lesional oral mucosa epithelium of lichen planus and hyperkeratosis were counted and statistically analysed. In the healthy oral mucosa, K20-positive Merkel cells with and without close association to the intraepithelial NF200-ir nerve fibres were detected. In the lesional oral mucosa of lichen planus and hyperkeratosis patients, extremely rare NF200-ir nerve fibres were detected only in the lamina propria. Compared with healthy tissues, lichen planus and hyperkeratosis tissues had significantly decreased numbers of NF200-ir nerve fibres in the oral mucosal epithelium. Lichen planus and hyperkeratosis were associated with the absence of Aβ-nerve endings in the oral mucosal epithelium. Thus, we conclude that mechanosensation mediated by the Merkel cell-neurite complex in the oral mucosal epithelium is impaired in lichen planus and hyperkeratosis.

  11. KANK1 inhibits cell growth by inducing apoptosis though regulating CXXC5 in human malignant peripheral nerve sheath tumors

    Science.gov (United States)

    Cui, Zhibin; Shen, Yingjia; Chen, Kenny H.; Mittal, Suresh K.; Yang, Jer-Yen; Zhang, GuangJun

    2017-01-01

    Malignant peripheral nerve sheath tumors (MPNSTs) are a type of rare sarcomas with a poor prognosis due to its highly invasive nature and limited treatment options. Currently there is no targeted-cancer therapy for this type of malignancy. Thus, it is important to identify more cancer driver genes that may serve as targets of cancer therapy. Through comparative oncogenomics, we have found that KANK1 was a candidate tumor suppressor gene (TSG) for human MPNSTs. Although KANK1 is known as a cytoskeleton regulator, its tumorigenic function in MPNSTs remains largely unknown. In this study, we report that restoration of KANK1 in human MPNST cells inhibits cell growth both in human cell culture and xenograft mice by increasing apoptosis. Consistently, knockdown of KANK1 in neurofibroma cells promoted cell growth. Using RNA-seq analysis, we identified CXXC5 and other apoptosis-related genes, and demonstrated that CXXC5 is regulated by KANK1. Knockdown of CXXC5 was found to diminish KANK1-induced apoptosis in MPNST cells. Thus, KANK1 inhibits MPNST cell growth though CXXC5 mediated apoptosis. Our results suggest that KANK1 may function as a tumor suppressor in human MPNSTs, and thus it may be useful for targeted therapy. PMID:28067315

  12. Multipotent stem cells isolated from the adult mouse retina are capable of producing functional photoreceptor cells.

    Science.gov (United States)

    Li, Tianqing; Lewallen, Michelle; Chen, Shuyi; Yu, Wei; Zhang, Nian; Xie, Ting

    2013-06-01

    Various stem cell types have been tested for their potential application in treating photoreceptor degenerative diseases, such as retinitis pigmentosa (RP) and age-related macular degeneration (AMD). Only embryonic stem cells (ESCs) have so far been shown to generate functional photoreceptor cells restoring light response of photoreceptor-deficient mice, but there is still some concern of tumor formation. In this study, we have successfully cultured Nestin(+)Sox2(+)Pax6(+) multipotent retinal stem cells (RSCs) from the adult mouse retina, which are capable of producing functional photoreceptor cells that restore the light response of photoreceptor-deficient rd1 mutant mice following transplantation. After they have been expanded for over 35 passages in the presence of FGF and EGF, the cultured RSCs still maintain stable proliferation and differentiation potential. Under proper differentiation conditions, they can differentiate into all the major retinal cell types found in the adult retina. More importantly, they can efficiently differentiate into photoreceptor cells under optimized differentiation conditions. Following transplantation into the subretinal space of slowly degenerating rd7 mutant eyes, RSC-derived photoreceptor cells integrate into the retina, morphologically resembling endogenous photoreceptors and forming synapases with resident retinal neurons. When transplanted into eyes of photoreceptor-deficient rd1 mutant mice, a RP model, RSC-derived photoreceptors can partially restore light response, indicating that those RSC-derived photoreceptors are functional. Finally, there is no evidence for tumor formation in the photoreceptor-transplanted eyes. Therefore, this study has demonstrated that RSCs isolated from the adult retina have the potential of producing functional photoreceptor cells that can potentially restore lost vision caused by loss of photoreceptor cells in RP and AMD.

  13. Multipotent stem cells isolated from the adult mouse retina are capable of producing functional photoreceptor cells

    Institute of Scientific and Technical Information of China (English)

    Tianqing Li; Michelle Lewallen; Shuyi Chen; Wei Yu; Nian Zhang; Ting Xie

    2013-01-01

    Various stem cell types have been tested for their potential application in treating photoreceptor degenerative diseases,such as retinitis pigmentosa (RP) and age-related macular degeneration (AMD).Only embryonic stem cells (ESCs) have so far been shown to generate functional photoreceptor cells restoring light response of photoreceptordeficient mice,but there is still some concern of tumor formation.In this study,we have successfully cultured Nestin+Sox2+Pax6+ multipotent retinal stem cells (RSCs) from the adult mouse retina,which are capable of producing functional photoreceptor cells that restore the light response of photoreceptor-deficient rd1 mutant mice following transplantation.After they have been expanded for over 35 passages in the presence of FGF and EGF,the cultured RSCs still maintain stable proliferation and differentiation potential.Under proper differentiation conditions,they can differentiate into all the major retinal cell types found in the adult retina.More importantly,they can efficiently differentiate into photoreceptor cells under optimized differentiation conditions.Following transplantation into the subretinal space of slowly degenerating rd7 mutant eyes,RSC-derived photoreceptor cells integrate into the retina,morphologically resembling endogenous photoreceptors and forming synapases with resident retinal neurons.When transplanted into eyes of photoreceptor-deficient rd1 mutant mice,a RP model,RSC-derived photoreceptors can partially restore light response,indicating that those RSC-derived photoreceptors are functional.Finally,there is no evidence for tumor formation in the photoreceptor-transplanted eyes.Therefore,this study has demonstrated that RSCs isolated from the adult retina have the potential of producing functional photoreceptor cells that can potentially restore lost vision caused by loss of photoreceptor cells in RP and AMD.

  14. Nasal-Type Extranodal Natural Killer/T-cell Neurolymphomatosis Confined to the Lumbar Nerve Roots: A Case Report

    Energy Technology Data Exchange (ETDEWEB)

    Park, Jong Chun; Mun, Sung Hee; Lee, Young Hwan [Catholic University, Daegu (Korea, Republic of)

    2009-11-15

    Neurolymphomatosis refers to lymphoma that has infiltrated the peripheral nervous system and this is the least common clinical presentation of nervous system lymphoma. Most neurolymphomatosis is due to B-cell non-Hodgkin lymphoma, and most patients show lymphomatous infiltration in the meninges and brain parenchyma, in addition to peripheral nervous system involvement. We diagnosed a case of neurolymphomatosis that was confined to the right 4th and 5th lumbar nerve roots without involvement of the meninges or brain parenchyma in a patient with the nasal-type extranodal natural killer/T-cell lymphoma. We made this diagnosis based on the MRI and 18F-FDG PET-CT findings and the clinical manifestations.

  15. Changes in the blood-nerve barrier after sciatic nerve cold injury: indications supporting early treatment

    Directory of Open Access Journals (Sweden)

    Hao Li

    2015-01-01

    Full Text Available Severe edema in the endoneurium can occur after non-freezing cold injury to the peripheral nerve, which suggests damage to the blood-nerve barrier. To determine the effects of cold injury on the blood-nerve barrier, the sciatic nerve on one side of Wistar rats was treated with low temperatures (3-5°C for 2 hours. The contralateral sciatic nerve was used as a control. We assessed changes in the nerves using Evans blue as a fluid tracer and morphological methods. Excess fluid was found in the endoneurium 1 day after cold injury, though the tight junctions between cells remained closed. From 3 to 5 days after the cold injury, the fluid was still present, but the tight junctions were open. Less tracer leakage was found from 3 to 5 days after the cold injury compared with 1 day after injury. The cold injury resulted in a breakdown of the blood-nerve barrier function, which caused endoneurial edema. However, during the early period, the breakdown of the blood-nerve barrier did not include the opening of tight junctions, but was due to other factors. Excessive fluid volume produced a large increase in the endoneurial fluid pressure, prevented liquid penetration into the endoneurium from the microvasculature. These results suggest that drug treatment to patients with cold injuries should be administered during the early period after injury because it may be more difficult for the drug to reach the injury site through the microcirculation after the tissue fluid pressure becomes elevated.

  16. Cloning of the Eukaryotic Expression Vector with Nerve Growth Factor in Rats and Its Effects on Proliferation and Differentiation of Mesencephal Neural Stem Cells of Fetal Rats

    Institute of Scientific and Technical Information of China (English)

    Minhua LIN; Lin YANG; Rong FU; Hongyang ZHAO

    2008-01-01

    Summary: The eukaryotic expression vector containing full-length cDNA sequence of rate nerve growth factor (NGF) β subunit was constructed and its effects on proliferation and differentiation of neural stem cells were observed. By using PCR, full-length cDNA sequence of NGF β subunit in rats was cloned and ligated into the eukaryotic expression vector pEGFP-N1-NGE The recombinant plasmid pEGFP-N1-NGF was transfected into the mesencephal neural stem cells of embryonic rats by Lipofectamin and transiently expressed. MTT method was used to determine the effects of NGF on proliferation of neural stem cells, and under phase-contrast microscopy, the effects of NGF on growth of nervous processes following differentiation of neural stem cells were observed. Sequence analysis indicated that the cloned full-length cDNA sequence of rat NGF β was identical to that of published sequence encoding NGF in gene GeneBank. The transfection of recombinant plasmid pEGFP-N1-NGF into mesencephal neural stem cells of embryonic rats could obviously promote proliferation of neural stem cells and faciliate the growth of neural stem cells-derived nerve cells. It was suggested that neural stem cells could be used as a vehicle of gene transfer, and the expression of NGF β subunit in the neural stem cells could promote the growth of nerve cells derived from neural stem cells.

  17. Cytokine-producing T cell subsets in human leishmaniasis

    DEFF Research Database (Denmark)

    Kemp, Kåre

    2000-01-01

    Leishmania specific Th1/Th2 cells have been identified in humans as well as in mice. There is a correlation between the clinical outcome of the infection and the cytokine response profile. Generally, the production of Th2 cytokines leads to severe infection, whereas the production of Th1 cytokine...... cells mutually down-regulate each other. However, the presence of antigen specific regulatory T cell subsets may provide an environment that allows the presence of both Th1 and Th2 cells....

  18. Distribution and localization of fibroblast growth factor-8 in rat brain and nerve cells during neural stem/progenitor cell differentiation

    Institute of Scientific and Technical Information of China (English)

    Jiang Lu; Dongsheng Li; Kehuan Lu

    2012-01-01

    The present study explored the distribution and localization of fibroblast growth factor-8 and its potential receptor,fibroblast growth factor receptor-3,in adult rat brain in vivo and in nerve cells during differentiation of neural stem/progenitor cells in vitro.Immunohistochemistry was used to examine the distribution of fibroblast growth factor-8 in adult rat brain in vivo.Localization of fibroblast growth factor-8 and fibroblast growth factor receptor-3 in cells during neural stem/progenitor cell differentiation in vitro was detected by immunofluorescence.Flow cytometry and immunofluorescence were used to evaluate the effect of an anti-fibroblast growth factor-8 antibody on neural stem/progenitor cell differentiation and expansion in vitro.Results from this study confirmed that fibroblast growth factor-8 was mainly distributed in adult midbrain,namely the substantia nigra,compact part,dorsal tier,substantia nigra and reticular part,but was not detected in the forebrain comprising the caudate putamen and striatum.Unusual results were obtained in retrosplenial locations of adult rat brain.We found that fibroblast growth factor-8 and fibroblast growth factor receptor-3 were distributed on the cell membrane and in the cytoplasm of nerve cells using immunohistochemistry and immunofluorescence analyses.We considered that the distribution of fibroblast growth factor-8 and fibroblast growth factor receptor-3 in neural cells corresponded to the characteristics of fibroblast growth factor-8,a secretory factor.Addition of an anti-fibroblast growth factor-8 antibody to cultures significantly affected the rate of expansion and differentiation of neural stem/progenitor cells.In contrast,addition of recombinant fibroblast growth factor-8 to differentiation medium promoted neural stem/progenitor cell differentiation and increased the final yields of dopaminergic neurons and total neurons.Our study may help delineate the important roles of fibroblast growth factor-8 in brain

  19. Increased encapsulated cell biodelivery of nerve growth factor in the brain by transposon-mediated gene transfer.

    Science.gov (United States)

    Fjord-Larsen, L; Kusk, P; Emerich, D F; Thanos, C; Torp, M; Bintz, B; Tornøe, J; Johnsen, A H; Wahlberg, L U

    2012-10-01

    Nerve growth factor (NGF) is a potential therapeutic agent for Alzheimer's disease (AD) as it has positive effects on the basal forebrain cholinergic neurons whose degeneration correlates with the cognitive decline in AD. We have previously described an encapsulated cell biodelivery device, NsG0202, capable of local delivery of NGF by a genetically modified human cell line, NGC-0295. The NsG0202 devices have shown promising safety and therapeutic results in a small phase 1b clinical study. However, results also show that the NGF dose could advantageously be increased. We have used the sleeping beauty transposon expression technology to establish a new clinical grade cell line, NGC0211, with at least 10 times higher NGF production than that of NGC-0295. To test whether encapsulation of this cell line provides a relevant dose escalation step in delivering NGF for treatment of the cognitive decline in AD patients, we have validated the bioactivity of devices with NGC0211 and NGC-0295 cells in normal rat striatum as well as in the quinolinic acid striatal lesion model. These preclinical animal studies show that implantation of devices with NGC0211 cells lead to significantly higher NGF output, which in both cases correlate with highly improved potency.

  20. Islet Brain 1 Protects Insulin Producing Cells against Lipotoxicity.

    Science.gov (United States)

    Brajkovic, Saška; Ferdaoussi, Mourad; Pawlowski, Valérie; Ezanno, Hélène; Plaisance, Valérie; Zmuda, Erik; Hai, Tsonwin; Annicotte, Jean-Sébastien; Waeber, Gérard; Abderrahmani, Amar

    2016-01-01

    Chronic intake of saturated free fatty acids is associated with diabetes and may contribute to the impairment of functional beta cell mass. Mitogen activated protein kinase 8 interacting protein 1 also called islet brain 1 (IB1) is a candidate gene for diabetes that is required for beta cell survival and glucose-induced insulin secretion (GSIS). In this study we investigated whether IB1 expression is required for preserving beta cell survival and function in response to palmitate. Chronic exposure of MIN6 and isolated rat islets cells to palmitate led to reduction of the IB1 mRNA and protein content. Diminution of IB1 mRNA and protein level relied on the inducible cAMP early repressor activity and proteasome-mediated degradation, respectively. Suppression of IB1 level mimicked the harmful effects of palmitate on the beta cell survival and GSIS. Conversely, ectopic expression of IB1 counteracted the deleterious effects of palmitate on the beta cell survival and insulin secretion. These findings highlight the importance in preserving the IB1 content for protecting beta cell against lipotoxicity in diabetes.

  1. Secretion of nerve growth factor, brain-derived neurotrophic factor, and glial cell-line derived neurotrophic factor in co-culture of four cell types in cerebrospinal fluid-containing medium

    Institute of Scientific and Technical Information of China (English)

    Sanjiang Feng; Minghua Zhuang; Rui Wu

    2012-01-01

    The present study co-cultured human embryonic olfactory ensheathing cells, human Schwann cells, human amniotic epithelial cells and human vascular endothelial cells in complete culture medium- containing cerebrospinal fluid. Enzyme linked immunosorbent assay was used to detect nerve growth factor, brain-derived neurotrophic factor, and glial cell line-derived neurotrophic factor secretion in the supernatant of co-cultured cells. Results showed that the number of all cell types reached a peak at 7–10 days, and the expression of nerve growth factor, brain-derived neurotrophic factor, and glial cell line-derived neurotrophic factor peaked at 9 days. Levels of secreted nerve growth factor were four-fold higher than brain-derived neurotrophic factor, which was three-fold higher than glial cell line-derived neurotrophic factor. Increasing concentrations of cerebrospinal fluid (10%, 20% and 30%) in the growth medium caused a decrease of neurotrophic factor secretion. Results indicated co-culture of human embryonic olfactory ensheathing cells, human Schwann cells, human amniotic epithelial cells and human vascular endothelial cells improved the expression of nerve growth factor, brain-derived neurotrophic factor, and glial cell line-derived neurotrophic factor. The reduction of cerebrospinal fluid extravasation at the transplant site after spinal cord injury is beneficial for the survival and secretion of neurotrophic factors from transplanted cells.

  2. Cloned calves produced by nuclear transfer from cultured cumulus cells

    Institute of Scientific and Technical Information of China (English)

    AN; Xiaorong(安晓荣); GOU; Kemian(苟克勉); ZHU; Shien(朱士恩); GUAN; Hong(关宏); HOU; Jian(侯健); LIN; Aixing(林爱星); ZENG; Shenming(曾申明); TIAN; Jianhui(田见辉); CHEN; Yongfu(陈永福)

    2002-01-01

    Short-term cultured cumulus cell lines (1-5BCC) derived from 5 individual cows were used in nuclear transfer (NT) and 1188 enucleated bovine oocytes matured in vitro were used as nuclear recipients. A total of 931 (78.4%) cloned embryos were reconstructed, of which 763 (82%) cleaved, 627 (67.3%) developed to 8-cell stage, and 275 (29.5%) reached blastocyst stage. The average cell number of blastocysts was 124±24.5 (n=20). In this study, the effects of donor cell sources, serum starvation of donor cells, time interval from fusion to activation (IFA) were also tested on cloning efficiency. These results showed that blastocyst rates of embryos reconstructed from 5 different individuals cells were significantly different among them (14.1%, 45.2%, 27.3%, 34.3%, vs 1.5%, P0.05); and that blastocyst rate (20.3%) of the group with fusion/activation interval of 2-3 h, was significantly lower than that of the 3-6 h groups (31.0%), while not significantly different among 3-4 h (P < 0.05), 4-5 h, and 5-6 h groups (P ≥ 0.05). Sixty-three thawed NT blastocysts were transferred to 31 recipient cows, of which 4 pregnancies were established and two cloned calves were given birth. These results indicate that serum starvation of cumulus cells is not a key factor for successful bovine cloning, while IFA treatment and sources of donor cells have effects on cloning efficiency.

  3. Producing fully ES cell-derived mice from eight-cell stage embryo injections.

    Science.gov (United States)

    DeChiara, Thomas M; Poueymirou, William T; Auerbach, Wojtek; Frendewey, David; Yancopoulos, George D; Valenzuela, David M

    2010-01-01

    In conventional methods for the generation of genetically modified mice, gene-targeted embryonic stem (ES) cells are injected into blastocyst-stage embryos or are aggregated with morula-stage embryos, which are then transferred to the uterus of a surrogate mother. F0 generation mice born from the embryos are chimeras composed of genetic contributions from both the modified ES cells and the recipient embryos. Obtaining a mouse strain that carries the gene-targeted mutation requires breeding the chimeras to transmit the ES cell genetic component through the germ line to the next (F1) generation (germ line transmission, GLT). To skip the chimera stage, we developed the VelociMouse method, in which injection of genetically modified ES cells into eight-cell embryos followed by maturation to the blastocyst stage and transfer to a surrogate mother produces F0 generation mice that are fully derived from the injected ES cells and exhibit a 100% GLT efficiency. The method is simple and flexible. Both male and female ES cells can be introduced into the eight-cell embryo by any method of injection or aggregation and using all ES cell and host embryo combinations from inbred, hybrid, and outbred genetic backgrounds. The VelociMouse method provides several unique opportunities for shortening project timelines and reducing mouse husbandry costs. First, as VelociMice exhibit 100% GLT, there is no need to test cross chimeras to establish GLT. Second, because the VelociMouse method permits efficient production of ES cell-derived mice from female ES cells, XO ES cell subclones, identified by screening for spontaneous loss of the Y chromosome, can be used to generate F0 females that can be bred with isogenic F0 males derived from the original targeted ES cell clone to obtain homozygous mutant mice in the F1 generation. Third, as VelociMice are genetically identical to the ES cells from which they were derived, the VelociMouse method opens up myriad possibilities for creating mice with

  4. Mechanism of sphingosine 1-phosphate- and lysophosphatidic Acid-induced up-regulation of adhesion molecules and eosinophil chemoattractant in nerve cells.

    LENUS (Irish Health Repository)

    Costello, Richard W

    2012-02-01

    The lysophospholipids sphingosine 1-phosphate (S1P) and lysophosphatidic acid (LPA) act via G-protein coupled receptors S1P(1-5) and LPA(1-3) respectively, and are implicated in allergy. Eosinophils accumulate at innervating cholinergic nerves in asthma and adhere to nerve cells via intercellular adhesion molecule-1 (ICAM-1). IMR-32 neuroblastoma cells were used as an in vitro cholinergic nerve cell model. The G(i) coupled receptors S1P(1), S1P(3), LPA(1), LPA(2) and LPA(3) were expressed on IMR-32 cells. Both S1P and LPA induced ERK phosphorylation and ERK- and G(i)-dependent up-regulation of ICAM-1 expression, with differing time courses. LPA also induced ERK- and G(i)-dependent up-regulation of the eosinophil chemoattractant, CCL-26. The eosinophil granule protein eosinophil peroxidase (EPO) induced ERK-dependent up-regulation of transcription of S1P(1), LPA(1), LPA(2) and LPA(3), providing the situation whereby eosinophil granule proteins may enhance S1P- and\\/or LPA- induced eosinophil accumulation at nerve cells in allergic conditions.

  5. Mechanism of sphingosine 1-phosphate- and lysophosphatidic Acid-induced up-regulation of adhesion molecules and eosinophil chemoattractant in nerve cells.

    LENUS (Irish Health Repository)

    Costello, Richard W

    2011-05-01

    The lysophospholipids sphingosine 1-phosphate (S1P) and lysophosphatidic acid (LPA) act via G-protein coupled receptors S1P(1-5) and LPA(1-3) respectively, and are implicated in allergy. Eosinophils accumulate at innervating cholinergic nerves in asthma and adhere to nerve cells via intercellular adhesion molecule-1 (ICAM-1). IMR-32 neuroblastoma cells were used as an in vitro cholinergic nerve cell model. The G(i) coupled receptors S1P(1), S1P(3), LPA(1), LPA(2) and LPA(3) were expressed on IMR-32 cells. Both S1P and LPA induced ERK phosphorylation and ERK- and G(i)-dependent up-regulation of ICAM-1 expression, with differing time courses. LPA also induced ERK- and G(i)-dependent up-regulation of the eosinophil chemoattractant, CCL-26. The eosinophil granule protein eosinophil peroxidase (EPO) induced ERK-dependent up-regulation of transcription of S1P(1), LPA(1), LPA(2) and LPA(3), providing the situation whereby eosinophil granule proteins may enhance S1P- and\\/or LPA- induced eosinophil accumulation at nerve cells in allergic conditions.

  6. [Structural anatomy of cranial nerves (V, VII, VIII, IX, X)].

    Science.gov (United States)

    Guclu, B; Meyronet, D; Simon, E; Streichenberger, N; Sindou, M; Mertens, P

    2009-04-01

    This study reports a review of the literature on the structural anatomy of the Vth, VIIth, VIIIth, IXth, and Xth cranial nerves, known to harbor dysfunction syndromes in humans. Because these dysfunctions are hypothesized to be caused by neurovascular conflicts at the root entry/exit zone and the transitional zone between central and peripheral myelinization, this investigation focused on the study and description of this junction. All the cranial nerves, except the optic and olfactory nerves, which are considered to be more a direct expansion of the central nervous system, have a transitional zone between central myelin (coming from oligodendrocytes) and peripheral myelin (produced by Schwann cells). The human studies reported in the literature argue in favor of a dome-shaped transitional zone directed to the periphery. It seems that this junctional region is situated more peripherally in sensory nerves than in motor nerves. The transitional zone is situated very peripherally for the cochlear and vestibular nerves, and on the contrary very close to its exit from the brain stem for the facial nerve.

  7. Dorsal root ganglia neurons and differentiated adipose-derived stem cells: an in vitro co-culture model to study peripheral nerve regeneration.

    Science.gov (United States)

    de Luca, Alba C; Faroni, Alessandro; Reid, Adam J

    2015-02-26

    Dorsal root ganglia (DRG) neurons, located in the intervertebral foramina of the spinal column, can be used to create an in vitro system facilitating the study of nerve regeneration and myelination. The glial cells of the peripheral nervous system, Schwann cells (SC), are key facilitators of these processes; it is therefore crucial that the interactions of these cellular components are studied together. Direct contact between DRG neurons and glial cells provides additional stimuli sensed by specific membrane receptors, further improving the neuronal response. SC release growth factors and proteins in the culture medium, which enhance neuron survival and stimulate neurite sprouting and extension. However, SC require long proliferation time to be used for tissue engineering applications and the sacrifice of an healthy nerve for their sourcing. Adipose-derived stem cells (ASC) differentiated into SC phenotype are a valid alternative to SC for the set-up of a co-culture model with DRG neurons to study nerve regeneration. The present work presents a detailed and reproducible step-by-step protocol to harvest both DRG neurons and ASC from adult rats; to differentiate ASC towards a SC phenotype; and combines the two cell types in a direct co-culture system to investigate the interplay between neurons and SC in the peripheral nervous system. This tool has great potential in the optimization of tissue-engineered constructs for peripheral nerve repair.

  8. A combination of gangliosides and nerve growth factor alleviates lipopolysaccharide-induced neuronal cells damage and its mechanism

    Directory of Open Access Journals (Sweden)

    Song Ying

    2017-01-01

    Full Text Available Objective: To evaluate the effect of gangliosides (GM1 in combination with nerve growth factor (NGF against neuronal cells damage evoked by lipopolysaccharide (LPS, and tries to uncover its probable mechanism. Methods: (1 Cell viability was measured using Methyl thiazolyl tetrazolium (MTT method, which was also determined the optimum concentration of LPS for the damage models; meanwhile, cell morphology was observed by microscope. (2 The expression level of NF-κB was detected by RT-PCR. (3 Finally, NF-κB inhibitor pyrollidine dithiocarbamate (PDTC was treated for the research of NF-κB pathway. Results: (1 MTT results shown that the LPS injury was dose-dependent, and 100nmol/L was selected as the optimum damage concentration. (2 Through the morphological observation, MTT and RT-PCR analysis, we found that GM1 and NGF both can protect cells against LPS injury; interestingly, combination of GM1 and NGF had a slighter LPS injury than GM1 administration alone. Moreover, the expression of NF-κB in combination group was lower than that in GM1 group, indicated that blockage of NF-κB pathway was better for cells living. Conclusion: Combination of GM1 and NGF has a better protective act on LPS injury than GM1 alone. The mechanism may have some connections with NF-κB pathways.

  9. Post-embryonic nerve-associated precursors to adult pigment cells: genetic requirements and dynamics of morphogenesis and differentiation.

    Directory of Open Access Journals (Sweden)

    Erine H Budi

    2011-05-01

    Full Text Available The pigment cells of vertebrates serve a variety of functions and generate a stunning variety of patterns. These cells are also implicated in human pathologies including melanoma. Whereas the events of pigment cell development have been studied extensively in the embryo, much less is known about morphogenesis and differentiation of these cells during post-embryonic stages. Previous studies of zebrafish revealed genetically distinct populations of embryonic and adult melanophores, the ectotherm homologue of amniote melanocytes. Here, we use molecular markers, vital labeling, time-lapse imaging, mutational analyses, and transgenesis to identify peripheral nerves as a niche for precursors to adult melanophores that subsequently migrate to the skin to form the adult pigment pattern. We further identify genetic requirements for establishing, maintaining, and recruiting precursors to the adult melanophore lineage and demonstrate novel compensatory behaviors during pattern regulation in mutant backgrounds. Finally, we show that distinct populations of latent precursors having differential regenerative capabilities persist into the adult. These findings provide a foundation for future studies of post-embryonic pigment cell precursors in development, evolution, and neoplasia.

  10. PDX1-engineered embryonic stem cell-derived insulin producing cells regulate hyperglycemia in diabetic mice

    Directory of Open Access Journals (Sweden)

    Raikwar Sudhanshu P

    2012-10-01

    Full Text Available Abstract Background Type 1 diabetes can be treated by the transplantation of cadaveric whole pancreata or isolated pancreatic islets. However, this form of treatment is hampered by the chronic shortage of cadaveric donors. Embryonic stem (ES cell-derived insulin producing cells (IPCs offer a potentially novel source of unlimited cells for transplantation to treat type 1 and possibly type 2 diabetes. However, thus far, the lack of a reliable protocol for efficient differentiation of ES cells into IPCs has hindered the clinical exploitation of these cells. Methods To efficiently generate IPCs using ES cells, we have developed a double transgenic ES cell line R1Pdx1AcGFP/RIP-Luc that constitutively expresses pancreatic β-cell-specific transcription factor pancreatic and duodenal homeobox gene 1 (Pdx1 as well as rat insulin promoter (RIP driven luciferase reporter. We have established several protocols for the reproducible differentiation of ES cells into IPCs. The differentiation of ES cells into IPCs was monitored by immunostaining as well as real-time quantitative RT-PCR for pancreatic β-cell-specific markers. Pancreatic β-cell specific RIP became transcriptionally active following the differentiation of ES cells into IPCs and induced the expression of the luciferase reporter. Glucose stimulated insulin secretion by the ES cell-derived IPCs was measured by ELISA. Further, we have investigated the therapeutic efficacy of ES cell-derived IPCs to correct hyperglycemia in syngeneic streptozotocin (STZ-treated diabetic mice. The long term fate of the transplanted IPCs co-expressing luciferase in syngeneic STZ-induced diabetic mice was monitored by real time noninvasive in vivo bioluminescence imaging (BLI. Results We have recently demonstrated that spontaneous in vivo differentiation of R1Pdx1AcGFP/RIP-Luc ES cell-derived pancreatic endoderm-like cells (PELCs into IPCs corrects hyperglycemia in diabetic mice. Here, we investigated whether R1Pdx1Ac

  11. 干细胞移植与神经修复★%Stem cell transplantation for nerve repair

    Institute of Scientific and Technical Information of China (English)

    孟祥鹏; 孙宝红; 陈立杰

    2013-01-01

    BACKGROUND: With development of life science, tissue engineering technology involving stem cells likely enhances the regenerative capacity of injured nerve tissue. OBJECTIVE: To review the clinical application of several important stem cells in repair of nervous system injury and to clarify their application range and clinical validity, providing experimental evidence for clinical physicians. METHODS: A computer-based online retrieval of PubMed database was performed for the related articles regarding stem cel transplantation for nerve repair published during 1998-2011, with the key words “stem cells;transplantation; nerve repair” in English. Meanwhile, relevant articles published in Wanfang database during 2006-2011 were retrieved with key words “ stem cells; transplantation; nerve repair” in Chinese. RESULTS AND CONCLUSION: Through the initial retrieval, 284 basic research or clinical application papers were searched. These papers were analyzed in terms of source, differentiation, characteristics of transplanted stem cells as wel as the effects of stem cel transplantation on nerve repair irrespective of humans or animals as subjects. After excluding papers with repetitive contents and case report papers, 28 papers were included in the final analysis. This suggests that stem cel transplantation provides a novel pathway to treatment of nervous system diseases and enables structure repair and functional reconstruction of nerve tissue that can be traditional y considered impossible. Stem cel transplantation is a brand-new research field and there are many problems to be solved.%  背景:随着生命科学发展,利用干细胞来源的组织工程技术,有可能使受损的神经组织恢复再生能力。目的:针对较为重要的几种干细胞在神经系统修复中的临床应用做一综述,以期阐明各自适用范围和临床效度,为临床医师按需选择提供参考。方法:应用计算机检索 PubMed 数据库中1998

  12. The Evaluation of Nerve Growth Factor Over Expression on Neural Lineage Specific Genes in Human Mesenchymal Stem Cells

    Directory of Open Access Journals (Sweden)

    Mortazavi Yousef

    2016-07-01

    Full Text Available Objective Treatment and repair of neurodegenerative diseases such as brain tumors, spinal cord injuries, and functional disorders, including Alzheimer’s disease, are challenging problems. A common treatment approach for such disorders involves the use of mesenchymal stem cells (MSCs as an alternative cell source to replace injured cells. However, use of these cells in hosts may potentially cause adverse outcomes such as tumorigenesis and uncontrolled differentiation. In attempt to generate mesenchymal derived neural cells, we have infected MSCs with recombinant lentiviruses that expressed nerve growth factor (NGF and assessed their neural lineage genes. Materials and Methods In this experimental study, we cloned the NGF gene sequence into a helper dependent lentiviral vector that contained the green fluorescent protein (GFP gene. The recombinant vector was amplified in DH5 bacterial cells. Recombinant viruses were generated in the human embryonic kidney 293 (HEK-293 packaging cell line with the helper vectors and analyzed under fluorescent microscopy. Bone marrow mesenchymal cells were infected by recombinant viruses for three days followed by assessment of neural differentiation. We evaluated expression of NGF through measurement of the NGF protein in culture medium by ELISA; neural specific genes were quantified by real-time polymerase chain reaction (PCR. Results We observed neural morphological changes after three days. Quantitative PCR showed that expressions of NESTIN, glial derived neurotrophic factor (GDNF, glial fibrillary acidic protein (GFAP and Microtubule-associated protein 2 (MAP2 genes increased following induction of NGF overexpression, whereas expressions of endogenous NGF and brain derived neural growth factor (BDNF genes reduced. Conclusion Ectopic expression of NGF can induce neurogenesis in MSCs. Direct injection of MSCs may cause tumorigenesis and an undesirable outcome. Therefore an alternative choice to overcome this

  13. The Hagfish Gland Thread Cell: A Fiber-Producing Cell Involved in Predator Defense

    Directory of Open Access Journals (Sweden)

    Douglas S. Fudge

    2016-05-01

    Full Text Available Fibers are ubiquitous in biology, and include tensile materials produced by specialized glands (such as silks, extracellular fibrils that reinforce exoskeletons and connective tissues (such as chitin and collagen, as well as intracellular filaments that make up the metazoan cytoskeleton (such as F-actin, microtubules, and intermediate filaments. Hagfish gland thread cells are unique in that they produce a high aspect ratio fiber from cytoskeletal building blocks within the confines of their cytoplasm. These threads are elaborately coiled into structures that readily unravel when they are ejected into seawater from the slime glands. In this review we summarize what is currently known about the structure and function of gland thread cells and we speculate about the mechanism that these cells use to produce a mechanically robust fiber that is almost one hundred thousand times longer than it is wide. We propose that a key feature of this mechanism involves the unidirectional rotation of the cell’s nucleus, which would serve to twist disorganized filaments into a coherent thread and impart a torsional stress on the thread that would both facilitate coiling and drive energetic unravelling in seawater.

  14. Methylcobalamin promotes the differentiation of Schwann cells and remyelination in lysophosphatidylcholine-induced demyelination of the rat sciatic nerve

    Directory of Open Access Journals (Sweden)

    Shunsuke eNishimoto

    2015-08-01

    Full Text Available Schwann cells (SCs are constituents of the peripheral nervous system. The differentiation of SCs in injured peripheral nerves is critical for regeneration after injury. Methylcobalamin (MeCbl is a vitamin B12 analog that is necessary for the maintenance of the peripheral nervous system. In this study, we estimated the effect of MeCbl on SCs. We showed that MeCbl downregulated the activity of Erk1/2 and promoted the expression of the myelin basic protein in SCs. In a dorsal root ganglion neuron–SC coculture system, myelination was promoted by MeCbl. In a focal demyelination rat model, MeCbl promoted remyelination and motor and sensory functional regeneration. MeCbl promoted the in vitro differentiation of SCs and in vivo myelination in a rat demyelination model and may be a novel therapy for several types of nervous disorders.

  15. An anatomical study of porcine peripheral nerve and its potential use in nerve tissue engineering

    Science.gov (United States)

    Zilic, Leyla; Garner, Philippa E; Yu, Tong; Roman, Sabiniano; Haycock, John W; Wilshaw, Stacy-Paul

    2015-01-01

    Current nerve tissue engineering applications are adopting xenogeneic nerve tissue as potential nerve grafts to help aid nerve regeneration. However, there is little literature that describes the exact location, anatomy and physiology of these nerves to highlight their potential as a donor graft. The aim of this study was to identify and characterise the structural and extracellular matrix (ECM) components of porcine peripheral nerves in the hind leg. Methods included the dissection of porcine nerves, localisation, characterisation and quantification of the ECM components and identification of nerve cells. Results showed a noticeable variance between porcine and rat nerve (a commonly studied species) in terms of fascicle number. The study also revealed that when porcine peripheral nerves branch, a decrease in fascicle number and size was evident. Porcine ECM and nerve fascicles were found to be predominately comprised of collagen together with glycosaminoglycans, laminin and fibronectin. Immunolabelling for nerve growth factor receptor p75 also revealed the localisation of Schwann cells around and inside the fascicles. In conclusion, it is shown that porcine peripheral nerves possess a microstructure similar to that found in rat, and is not dissimilar to human. This finding could extend to the suggestion that due to the similarities in anatomy to human nerve, porcine nerves may have utility as a nerve graft providing guidance and support to regenerating axons. PMID:26200940

  16. An anatomical study of porcine peripheral nerve and its potential use in nerve tissue engineering.

    Science.gov (United States)

    Zilic, Leyla; Garner, Philippa E; Yu, Tong; Roman, Sabiniano; Haycock, John W; Wilshaw, Stacy-Paul

    2015-09-01

    Current nerve tissue engineering applications are adopting xenogeneic nerve tissue as potential nerve grafts to help aid nerve regeneration. However, there is little literature that describes the exact location, anatomy and physiology of these nerves to highlight their potential as a donor graft. The aim of this study was to identify and characterise the structural and extracellular matrix (ECM) components of porcine peripheral nerves in the hind leg. Methods included the dissection of porcine nerves, localisation, characterisation and quantification of the ECM components and identification of nerve cells. Results showed a noticeable variance between porcine and rat nerve (a commonly studied species) in terms of fascicle number. The study also revealed that when porcine peripheral nerves branch, a decrease in fascicle number and size was evident. Porcine ECM and nerve fascicles were found to be predominately comprised of collagen together with glycosaminoglycans, laminin and fibronectin. Immunolabelling for nerve growth factor receptor p75 also revealed the localisation of Schwann cells around and inside the fascicles. In conclusion, it is shown that porcine peripheral nerves possess a microstructure similar to that found in rat, and is not dissimilar to human. This finding could extend to the suggestion that due to the similarities in anatomy to human nerve, porcine nerves may have utility as a nerve graft providing guidance and support to regenerating axons.

  17. Integrating Wind And Solar With Hydrogen Producing Fuel Cells

    NARCIS (Netherlands)

    Hemmes, K.

    2007-01-01

    The often proposed solution for the fluctuating wind energy supply is the conversion of the surplus of wind energy into hydrogen by means of electrolysis. In this paper a patented alternative is proposed consisting of the integration of wind turbines with internal reforming fuel-cells, capable of co

  18. Calcium Signaling in Mitral Cell Dendrites of Olfactory Bulbs of Neonatal Rats and Mice during Olfactory Nerve Stimulation and Beta-Adrenoceptor Activation

    Science.gov (United States)

    Yuan, Qi; Mutoh, Hiroki; Debarbieux, Franck; Knopfel, Thomas

    2004-01-01

    Synapses formed by the olfactory nerve (ON) provide the source of excitatory synaptic input onto mitral cells (MC) in the olfactory bulb. These synapses, which relay odor-specific inputs, are confined to the distally tufted single primary dendrites of MCs, the first stage of central olfactory processing. Beta-adrenergic modulation of electrical…

  19. Gene probes to detect cross-culture contamination in hormone producing cell lines

    DEFF Research Database (Denmark)

    Matsuba, I; Lernmark, A; Madsen, Ole Dragsbæk;

    1988-01-01

    Cross-culture contamination of cell lines propagated in continuous culture is a frequent event and particularly difficult to resolve in cells expressing similar phenotypes. We demonstrate that DNA-DNA hybridization to blotted endonuclease-digested cell DNA effectively detects cross-culture...... the effective use of gene probes to control the origin of cell cultures....... contamination to monitor inter-species as well as intra-species cross contamination. An insulin-producing cell-line, Clone-16, originally cloned from a human fetal endocrine pancreatic cell line did not produce human c-peptide as anticipated. DNA from these cells showed no hybridization to the human ALU...

  20. The Effect of Electrospun Gelatin Fibers Alignment on Schwann Cell and Axon Behavior and Organization in the Perspective of Artificial Nerve Design.

    Science.gov (United States)

    Gnavi, Sara; Fornasari, Benedetta Elena; Tonda-Turo, Chiara; Laurano, Rossella; Zanetti, Marco; Ciardelli, Gianluca; Geuna, Stefano

    2015-01-01

    Electrospun fibrous substrates mimicking extracellular matrices can be prepared by electrospinning, yielding aligned fibrous matrices as internal fillers to manufacture artificial nerves. Gelatin aligned nano-fibers were prepared by electrospinning after tuning the collector rotation speed. The effect of alignment on cell adhesion and proliferation was tested in vitro using primary cultures, the Schwann cell line, RT4-D6P2T, and the sensory neuron-like cell line, 50B11. Cell adhesion and proliferation were assessed by quantifying at several time-points. Aligned nano-fibers reduced adhesion and proliferation rate compared with random fibers. Schwann cell morphology and organization were investigated by immunostaining of the cytoskeleton. Cells were elongated with their longitudinal body parallel to the aligned fibers. B5011 neuron-like cells were aligned and had parallel axon growth when cultured on the aligned gelatin fibers. The data show that the alignment of electrospun gelatin fibers can modulate Schwann cells and axon organization in vitro, suggesting that this substrate shows promise as an internal filler for the design of artificial nerves for peripheral nerve reconstruction.

  1. 神经干细胞的迁移和网络化%Migration and nerve net building of neural stem cells

    Institute of Scientific and Technical Information of China (English)

    薛杉; 张旺明; 徐如祥

    2005-01-01

    目的:从神经干细胞的迁移现象及其定向迁移的可能机制,神经网络的研究进展和神经干细胞迁移和网络化的意义等方面进行综述.资料来源:应用计算机检索Pubmed数据库1970/2004期间的相关文章,限定文章语言种类为English,检索词为"neural stem cell,migration,nerve net".资料选择:对资料进行初审,选取神经干细胞迁移和网络化的随机和非随机对照实验.排除综述类和重复的文章.资料提炼:共收集到相关文献39篇,共检出16篇文献涉及神经干细胞的迁移和网络化研究的进展,其余文献均被排除.资料综合:对于神经系统退行性病变及严重损伤,神经干细胞移植有可能替代衰老变性和死亡的神经细胞,重建神经网络,恢复失去的脑功能.神经细胞的迁移和网络化原理可以应用以解决神经干细胞移植后的存活、分化、迁移及神经网络建立的问题,实现脑功能修复重建.结论:神经干细胞在特定的情况下可以迁移到预计的地方并建立神经网络.%OBJECTIVE: A series of recent studies have demonstrated the mechanism of migration and nerve net of neural stem cells. These theories have further substantiated neural stem cell transplantation. In view of these new findings, this paper reviewed the mode of migration and information of network. The significance of these theories was discussed.DATA SOURCE: We search on Pubmed with the key words "neural stem cell", "migration", and "nerve net", limiting the language to English and publication date from 1970 to 2004. At the same time we searched on CNKI.STUDY SELECTION: We selected the randomized and non-randomized controlled studies related to migration and nerve net building of neural stem cells. Review articles and articles with repetitive studies were excluded.DATA EXTRACTION: Among 39 papers selected, 16 papers concerning the development of this topic were selected, and the others were excluded.DATA SYNTHESIS: For

  2. Progress in amorphous silicon solar cells produced by reactive sputtering

    Science.gov (United States)

    Moustakas, T. D.

    The photovoltaic properties of reactively sputtered amorphous silicon are reviewed and it is shown that efficient PIN solar cells can be fabricated by the method of sputtering. The photovoltaic properties of the intrinsic films correlate with their structural and compositional inhomogeneities. Hydrogen incorporation and small levels of phosphorus and boron impurities also affect the photovoltaic properties through reduction of residual dangling bond related defects and modification of their occupation. The optical and transport properties of the doped P and N-films were found to depend sensitively on the amount of hydrogen and boron or phosphorus incorporation into the films as well as on their degree of crystallinity. Combination of the best intrinsic and doped films leads to PIN solar cell structures generating J(sc) of 13 mA/sq cm and V(oc) of between 0.85 to 0.95 volts. The efficiency of these devices, 5 to 6 percent, is limited by the low FF, typically about 50 percent. As a further test to the potential of this technology efficient tandem solar cell structures were fabricated, and device design concepts, such as the incorporation of optically reflective back contacts were tested.

  3. Neovibsanin B increases extracellular matrix proteins in optic nerve head cells via activation of Smad signalling pathway.

    Science.gov (United States)

    Wang, Zhen; Xu, Wei; Rong, Ao; Lin, Yan; Qiu, Xu-Ling; Qu, Shen; Lan, Xian-Hai

    2015-01-01

    The present study demonstrates the effect of neovibsanin B on the synthesis and deposition of ECM proteins and the signalling pathways used in optic nerve head (ONH) astrocytes and lamina cribrosa (LC) cells. For investigation of the signalling pathway used by neovibsanin B, ONH cells were treated with neovibsanin B. Western blot and immunostaining analyses were used to examine the phosphorylation of proteins involved in Smad and non-Smad signalling pathway. The results revealed that ONH cells on treatment with neovibsanin B showed enhanced synthesis of extracellular matrix (ECM) proteins. Neovibsanin B induced phosphorylation of canonical signalling proteins, Smad2/3. However phosphorylation of non-canonical signalling proteins, extracellular signal-regulated kinases, p38, and c-Jun N-terminal kinases (JNK) 1/2 remained unaffected. There was also increase in co-localization of pSmad2/3 with Co-Smad4 in the nucleus of ONH astrocytes and LC cells indicating activation of the canonical Smad signalling pathway. Treatment of ONH cells with SIS3, inhibitor of Smad3 phosphorylation reversed the neovibsanin B stimulated ECM expression as well as activation of canonical pathway signalling molecules. In addition, inhibition of Smad2 or Smad3 using small interfering RNA (siRNA) also suppressed neovibsanin B stimulated ECM protein synthesis in ONH astrocytes and LC cells. Thus neovibsanin B utilizes the canonical Smad signalling pathway to stimulate ECM synthesis in human ONH cells. The neovibsanin B induced ECM synthesis and activation of the canonical Smad signalling pathway may be due to its effect on transforming growth factor-β2 (TGF-β2). However, further studies are under process to understand the mechanism.

  4. Expression profiling and Ingenuity biological function analyses of interleukin-6- versus nerve growth factor-stimulated PC12 cells

    Directory of Open Access Journals (Sweden)

    Dimitriades-Schmutz Beatrice

    2009-02-01

    Full Text Available Abstract Background The major goal of the study was to compare the genetic programs utilized by the neuropoietic cytokine Interleukin-6 (IL-6 and the neurotrophin (NT Nerve Growth Factor (NGF for neuronal differentiation. Results The designer cytokine Hyper-IL-6 in which IL-6 is covalently linked to its soluble receptor s-IL-6R as well as NGF were used to stimulate PC12 cells for 24 hours. Changes in gene expression levels were monitored using Affymetrix GeneChip technology. We found different expression for 130 genes in IL-6- and 102 genes in NGF-treated PC12 cells as compared to unstimulated controls. The gene set shared by both stimuli comprises only 16 genes. A key step is upregulation of growth factors and functionally related external molecules known to play important roles in neuronal differentiation. In particular, IL-6 enhances gene expression of regenerating islet-derived 3 alpha (REG3A; 1084-fold, regenerating islet-derived 3 beta (REG3B/PAPI; 672-fold, growth differentiation factor 15 (GDF15; 80-fold, platelet-derived growth factor alpha (PDGFA; 69-fold, growth hormone releasing hormone (GHRH; 30-fold, adenylate cyclase activating polypeptide (PACAP; 20-fold and hepatocyte growth factor (HGF; 5-fold. NGF recruits GDF15 (131-fold, transforming growth factor beta 1 (TGFB1; 101-fold and brain-derived neurotrophic factor (BDNF; 89-fold. Both stimuli activate growth-associated protein 43 (GAP-43 indicating that PC12 cells undergo substantial neuronal differentiation. Moreover, IL-6 activates the transcription factors retinoic acid receptor alpha (RARA; 20-fold and early growth response 1 (Egr1/Zif268; 3-fold known to play key roles in neuronal differentiation. Ingenuity biological function analysis revealed that completely different repertoires of molecules are recruited to exert the same biological functions in neuronal differentiation. Major sub-categories include cellular growth and differentiation, cell migration, chemotaxis, cell

  5. Application of dielectric spectroscopy for monitoring high cell density in monoclonal antibody producing CHO cell cultivations.

    Science.gov (United States)

    Párta, László; Zalai, Dénes; Borbély, Sándor; Putics, Akos

    2014-02-01

    The application of dielectric spectroscopy was frequently investigated as an on-line cell culture monitoring tool; however, it still requires supportive data and experience in order to become a robust technique. In this study, dielectric spectroscopy was used to predict viable cell density (VCD) at industrially relevant high levels in concentrated fed-batch culture of Chinese hamster ovary cells producing a monoclonal antibody for pharmaceutical purposes. For on-line dielectric spectroscopy measurements, capacitance was scanned within a wide range of frequency values (100-19,490 kHz) in six parallel cell cultivation batches. Prior to detailed mathematical analysis of the collected data, principal component analysis (PCA) was applied to compare dielectric behavior of the cultivations. PCA analysis resulted in detecting measurement disturbances. By using the measured spectroscopic data, partial least squares regression (PLS), Cole-Cole, and linear modeling were applied and compared in order to predict VCD. The Cole-Cole and the PLS model provided reliable prediction over the entire cultivation including both the early and decline phases of cell growth, while the linear model failed to estimate VCD in the later, declining cultivation phase. In regards to the measurement error sensitivity, remarkable differences were shown among PLS, Cole-Cole, and linear modeling. VCD prediction accuracy could be improved in the runs with measurement disturbances by first derivative pre-treatment in PLS and by parameter optimization of the Cole-Cole modeling.

  6. Schwann cell-specific JAM-C-deficient mice reveal novel expression and functions for JAM-C in peripheral nerves.

    Science.gov (United States)

    Colom, Bartomeu; Poitelon, Yannick; Huang, Wenlong; Woodfin, Abigail; Averill, Sharon; Del Carro, Ubaldo; Zambroni, Desirée; Brain, Susan D; Perretti, Mauro; Ahluwalia, Amrita; Priestley, John V; Chavakis, Triantafyllos; Imhof, Beat A; Feltri, M Laura; Nourshargh, Sussan

    2012-03-01

    Junctional adhesion molecule-C (JAM-C) is an adhesion molecule expressed at junctions between adjacent endothelial and epithelial cells and implicated in multiple inflammatory and vascular responses. In addition, we recently reported on the expression of JAM-C in Schwann cells (SCs) and its importance for the integrity and function of peripheral nerves. To investigate the role of JAM-C in neuronal functions further, mice with a specific deletion of JAM-C in SCs (JAM-C SC KO) were generated. Compared to wild-type (WT) controls, JAM-C SC KO mice showed electrophysiological defects, muscular weakness, and hypersensitivity to mechanical stimuli. In addressing the underlying cause of these defects, nerves from JAM-C SC KO mice were found to have morphological defects in the paranodal region, exhibiting increased nodal length as compared to WTs. The study also reports on previously undetected expressions of JAM-C, namely on perineural cells, and in line with nociception defects of the JAM-C SC KO animals, on finely myelinated sensory nerve fibers. Collectively, the generation and characterization of JAM-C SC KO mice has provided unequivocal evidence for the involvement of SC JAM-C in the fine organization of peripheral nerves and in modulating multiple neuronal responses.

  7. Maslinic Acid Protected PC12 Cells Differentiated by Nerve Growth Factor against β-Amyloid-Induced Apoptosis.

    Science.gov (United States)

    Yang, Yu-wan; Tsai, Chia-wen; Mong, Mei-chin; Yin, Mei-chin

    2015-12-01

    β-Amyloid peptide (Abeta) was used to induce apoptosis in PC12 cells differentiated by nerve growth factor, and the protective activities of maslinic acid (MA) at 2-16 μM were examined. Abeta treatment lowered Bcl-2 expression, raised Bax expression, and decreased cell viability. MA pretreatments decreased Bax expression, raised the Bcl-2/Bax ratio, and increased cell viability. MA pretreatments retained glutathione content and decreased subsequent Abeta-induced release of reactive oxygen species, tumor necrosis factor-α, interleukin (IL)-1β, and IL-6. Abeta treatment up-regulated protein expression of p47(phox), gp91(phox), mitogen-activated protein kinase, advanced glycation end product receptor (RAGE), and nuclear factor-κ B (NF-κB). MA pretreatments at 2-16 μM suppressed the expression of proteins including gp91(phox), p47(phox), p-p38, and NF-κB p65, at 4-16 μM down-regulated RAGE and NF-κB p50 expression, and at 8 and 16 μM reduced p-ERK1/2 expression. These novel findings suggest that maslinic acid is a potent compound against Abeta-induced cytotoxicity.

  8. Effects of Co-grafts Mesenchymal Stem Cells and Nerve Growth Factor Suspension in the Repair of Spinal Cord Injury

    Institute of Scientific and Technical Information of China (English)

    FANG Huang; WANG Junfang; CHEN Anmin

    2006-01-01

    To investigate effect of the transplantation of mesenchymal stem cells (MSCs) in combination with nerve growth factor (NGF) on the repair of spinal cord injury (SCI) in adult rats, spinal cord of adult rats (n= 32) was injured by using the modified Allen' s method. One week after the injury, the injured cords were injected with Dubecco-modified Eagles medium (DMEM , Group Ⅰ), MSCs (Group Ⅱ), NGF (Group Ⅲ), and MSCs plus NGF (Group Ⅳ). One month and two months after the injury, rats were sacrificed and their injured cord tissues were sectioned for the identification of the transplanted cells. The axonal regeneration and the differentiation of MSCs were examined by immunocytochemical staining. At the same time, rats were subjected to behavioral tests by using the open-field BBB scoring system. Immunocytochemical staining showed that axonal regeneration and the transplanted cells partially expressed neuron-specific nuclear protein (NeuN) and glial fibrillary acidic protein (GFAP). At the same time, significant improvement in BBB locomotor rating scale (P<0.05) were observed in the treatment group. More importantly, further functional improvement were noted in the combined treatment group. MSCs could differentiate into neurons and astrocytes. MSCs and NGF can promote axonal regeneration and improve functional recovery. There might exist a synergistic effect between MSCs and NGF.

  9. Comparison of the effects of cumene hydroperoxide and hydrogen peroxide on Retzius nerve cells of the leech Haemopis sanguisuga.

    Science.gov (United States)

    Jovanovic, Zorica; Jovanovic, Svetlana

    2013-01-01

    Oxidative stress and the production of reactive oxygen species are known to play a major role in neuronal cell damage, but the exact mechanisms responsible for neuronal injury and death remain uncertain. In the present study, we examined the effects of oxidative stress on spontaneous spike activity and depolarizing outward potassium current by exposing the Retzius neurons of the leech to cumene hydroperoxide (CHP) and hydrogen peroxide (H(2)O(2)), the oxidants commonly used to examine oxidative mechanisms mediating cell death. We observed that relatively low concentrations of CHP (0.25, 1, and 1.5 mM) led to a marked prolongation of spontaneous repetitive activity. The prolonged action potentials showed an initial, spike-like depolarization followed by a plateau phase. In contrast, H(2)O(2) at the same and much higher concentrations (0.25 to 5 mM) did not significantly change the duration of spontaneous spike potentials of leech Retzius nerve cells (LRNCs). In the voltage clamp experiments, calcium-activated outward potassium currents, needed for the repolarization of the action potential, were suppressed with CHP, but not with H(2)O(2). The present findings indicate that CHP is a more potent oxidant and neurotoxin than H(2)O(2) and that the effect of CHP on the electrophysiological properties of LRNCs may be due to the inhibition of the potassium channels.

  10. Bilateral Facial Nerve Palsy in Acute B Cell Lymphoblastic Leukemia: A Case Report and Review of the Literature.

    Science.gov (United States)

    Sen, Shiraj; Gupta, Arjun; Friedman, Paul; Naina, Harris V

    2016-06-01

    Acute lymphoblastic leukemia (ALL) is a haematological malignancy that can involve the central nervous system (CNS). Less than 10 % of patients with ALL have CNS involvement at presentation. The cranial nerve most commonly affected is cranial nerve VII although bilateral involvement is rare. Management and outcomes of these patients are not well understood. Moreover bilateral Bells palsy as a presenting symptom of ALL is extremely uncommon. We report a very unusual presentation of ALL with bilateral facial nerve palsy, and discuss the management strategies and outcomes for patients with ALL that present with cranial nerve palsies.

  11. Evaluation of the retinal nerve fibre layer and ganglion cell complex thickness in pituitary macroadenomas without optic chiasmal compression

    Science.gov (United States)

    Cennamo, G; Auriemma, R S; Cardone, D; Grasso, L F S; Velotti, N; Simeoli, C; Di Somma, C; Pivonello, R; Colao, A; de Crecchio, G

    2015-01-01

    Purpose The aim of this prospective study was to measure the thickness of the circumpapillary retinal nerve fibre layer (cpRNFL) and the ganglion cell complex (GCC) using spectral domain optical coherence tomography (SD-OCT) in a cohort of consecutive de novo patients with pituitary macroadenomas without chiasmal compression. Patients and methods Twenty-two consecutive patients with pituitary macroadenoma without chiasmal compression (16 men, 6 women, aged 45.2±14.6 years, 43 eyes) entered the study between September 2011 and June 2013. Among them, 31.8% harboured a growth hormone-secreting pituitary adenoma, 27.3% a prolactin-secreting pituitary adenoma, 27.3% a corticotrophin-secreting pituitary adenoma, and 13.6% a non-secreting pituitary tumour. Eighteen subjects (nine females, nine males, mean age 36.47±6.37 years; 33 eyes) without pituitary adenoma, with normal ophthalmic examination, served as controls. In both patients and controls, cpRNFL and GCC thicknesses were measured by SD-OCT. Results Patients were significantly older (P=0.02) than controls. Best corrected visual acuity, intraocular pressure, colour fundus photography, and automatic perimetry test were within the normal range in patients and controls. Conversely, cpRNFL (P=0.009) and GCC (P<0.0001) were significantly thinner in patients than in controls. The average GCC (r=0.306, P=0.046) significantly correlated with the presence of arterial hypertension. OCT parameters did not differ significantly between patients with a tumour volume above the median and those with a tumour volume below the median. Conclusion Pituitary macroadenomas, even in the absence of chiasmal compression, may induce GCC and retinal nerve fibre layer thinning. SD-OCT may have a role in the early diagnosis and management of patients with pituitary tumours. PMID:25853400

  12. Electrochemically Produced Graphene for Microporous Layers in Fuel Cells.

    Science.gov (United States)

    Najafabadi, Amin Taheri; Leeuwner, Magrieta J; Wilkinson, David P; Gyenge, Előd L

    2016-07-01

    The microporous layer (MPL) is a key cathodic component in proton exchange membrane fuel cells owing to its beneficial influence on two-phase mass transfer. However, its performance is highly dependent on material properties such as morphology, porous structure, and electrical resistance. To improve water management and performance, electrochemically exfoliated graphene (EGN) microsheets are considered as an alternative to the conventional carbon black (CB) MPLs. The EGN-based MPLs decrease the kinetic overpotential and the Ohmic potential loss, whereas the addition of CB to form a composite EGN+CB MPL improves the mass-transport limiting current density drastically. This is reflected by increases of approximately 30 and 70 % in peak power densities at 100 % relative humidity (RH) compared with those for CB- and EGN-only MPLs, respectively. The composite EGN+CB MPL also retains the superior performance at a cathode RH of 20 %, whereas the CB MPL shows significant performance loss.

  13. Interleukin 1 expression is inducible by nerve growth factor in PC12 pheochromocytoma cells.

    OpenAIRE

    Alheim, K; Andersson, C; Tingsborg, S; Ziolkowska, M; Schultzberg, M. (Marianne); Bartfai, T

    1991-01-01

    Expression of the cytokine interleukin 1 alpha (IL-1 alpha) was demonstrated in the rat PC12 pheochromocytoma cell line by (i) immunohistochemistry using rabbit polyclonal antisera raised against the recombinant murine IL-1 alpha, (ii) an ELISA, and (iii) a specific cell conversion bioassay based on the use of LBRM33-1A5 cells. IL-1 alpha mRNA was demonstrated in the PC12 cells, by PCR amplification. Constitutive expression of IL-1 alpha in PC12 cells was demonstrated in all experiments, alth...

  14. Involvement of TRPM2 in peripheral nerve injury-induced infiltration of peripheral immune cells into the spinal cord in mouse neuropathic pain model.

    Directory of Open Access Journals (Sweden)

    Kouichi Isami

    Full Text Available Recent evidence suggests that transient receptor potential melastatin 2 (TRPM2 expressed in immune cells plays an important role in immune and inflammatory responses. We recently reported that TRPM2 expressed in macrophages and spinal microglia contributes to the pathogenesis of inflammatory and neuropathic pain aggravating peripheral and central pronociceptive inflammatory responses in mice. To further elucidate the contribution of TRPM2 expressed by peripheral immune cells to neuropathic pain, we examined the development of peripheral nerve injury-induced neuropathic pain and the infiltration of immune cells (particularly macrophages into the injured nerve and spinal cord by using bone marrow (BM chimeric mice by crossing wildtype (WT and TRPM2-knockout (TRPM2-KO mice. Four types of BM chimeric mice were prepared, in which irradiated WT or TRPM2-KO recipient mice were transplanted with either WT-or TRPM2-KO donor mouse-derived green fluorescence protein-positive (GFP(+ BM cells (TRPM2(BM+/Rec+, TRPM2(BM-/Rec+, TRPM2(BM+/Rec-, and TRPM2(BM-/Rec- mice. Mechanical allodynia induced by partial sciatic nerve ligation observed in TRPM2(BM+/Rec+ mice was attenuated in TRPM2(BM-/Rec+, TRPM2(BM+/Rec-, and TRPM2(BM-/Rec- mice. The numbers of GFP(+ BM-derived cells and Iba1/GFP double-positive macrophages in the injured sciatic nerve did not differ among chimeric mice 14 days after the nerve injury. In the spinal cord, the number of GFP(+ BM-derived cells, particularly GFP/Iba1 double-positive macrophages, was significantly decreased in the three TRPM2-KO chimeric mouse groups compared with TRPM2(BM+/Rec+ mice. However, the numbers of GFP(-/Iba1(+ resident microglia did not differ among chimeric mice. These results suggest that TRPM2 plays an important role in the infiltration of peripheral immune cells, particularly macrophages, into the spinal cord, rather than the infiltration of peripheral immune cells into the injured nerves and activation of spinal

  15. Ion channel noise can explain firing correlation in auditory nerves.

    Science.gov (United States)

    Moezzi, Bahar; Iannella, Nicolangelo; McDonnell, Mark D

    2016-10-01

    Neural spike trains are commonly characterized as a Poisson point process. However, the Poisson assumption is a poor model for spiking in auditory nerve fibres because it is known that interspike intervals display positive correlation over long time scales and negative correlation over shorter time scales. We have therefore developed a biophysical model based on the well-known Meddis model of the peripheral auditory system, to produce simulated auditory nerve fibre spiking statistics that more closely match the firing correlations observed in empirical data. We achieve this by introducing biophysically realistic ion channel noise to an inner hair cell membrane potential model that includes fractal fast potassium channels and deterministic slow potassium channels. We succeed in producing simulated spike train statistics that match empirically observed firing correlations. Our model thus replicates macro-scale stochastic spiking statistics in the auditory nerve fibres due to modeling stochasticity at the micro-scale of potassium channels.

  16. Polarized Th2 like cells, in the absence of Th0 cells, are responsible for lymphocyte produced IL-4 in high IgE-producer schistosomiasis patients

    Directory of Open Access Journals (Sweden)

    Soares-Silveira Alda

    2002-07-01

    Full Text Available Abstract Background Human resistance to re-infection with S. mansoni is correlated with high levels of anti-soluble adult worm antigens (SWAP IgE. Although it has been shown that IL-4 and IL-5 are crucial in establishing IgE responses in vitro, the active in vivo production of these cytokines by T cells, and the degree of polarization of Th2 vs. Th0 in human schistosomiasis is not known. To address this question, we determined the frequency of IL-4 and IFN-γ or IL-5 and IL-2 producing lymphocytes from schistosomiasis patients with high or low levels of IgE anti-SWAP. Results Our analysis showed that high and low IgE-producers responded equally to schistosomiasis antigens as determined by proliferation. Moreover, patients from both groups displayed similar percentages of circulating lymphocytes. However, high IgE-producers had an increased percentage of activated CD4+ T cells as compared to the low IgE-producers. Moreover, intracellular cytokine analysis, after short-term stimulation with anti-CD3/CD28 mAbs, showed that IgE high-producers display an increase in the percentage of T lymphocytes expressing IL-4 and IL-5 as compared to IgE low-responders. A coordinate control of the frequency of IL-4 and IL-5 producing lymphocytes in IgE high, but not IgE low-responders, was observed. Conclusions High IgE phenotype human schistosomiasis patients exhibit a coordinate regulation of IL-4 and IL-5 producing cells and the lymphocyte derived IL-4 comes from true polarized Th2 like cells, in the absence of measurable Th0 cells as measured by co-production of IL-4 and IFN-γ.

  17. BDNF Increases Survival and Neuronal Differentiation of Human Neural Precursor Cells Cotransplanted with a Nanofiber Gel to the Auditory Nerve in a Rat Model of Neuronal Damage

    Directory of Open Access Journals (Sweden)

    Yu Jiao

    2014-01-01

    Full Text Available Objectives. To study possible nerve regeneration of a damaged auditory nerve by the use of stem cell transplantation. Methods. We transplanted HNPCs to the rat AN trunk by the internal auditory meatus (IAM. Furthermore, we studied if addition of BDNF affects survival and phenotypic differentiation of the grafted HNPCs. A bioactive nanofiber gel (PA gel, in selected groups mixed with BDNF, was applied close to the implanted cells. Before transplantation, all rats had been deafened by a round window niche application of β-bungarotoxin. This neurotoxin causes a selective toxic destruction of the AN while keeping the hair cells intact. Results. Overall, HNPCs survived well for up to six weeks in all groups. However, transplants receiving the BDNF-containing PA gel demonstrated significantly higher numbers of HNPCs and neuronal differentiation. At six weeks, a majority of the HNPCs had migrated into the brain stem and differentiated. Differentiated human cells as well as neurites were observed in the vicinity of the cochlear nucleus. Conclusion. Our results indicate that human neural precursor cells (HNPC integration with host tissue benefits from additional brain derived neurotrophic factor (BDNF treatment and that these cells appear to be good candidates for further regenerative studies on the auditory nerve (AN.

  18. [Application of induced pluripotent stem (iPS) cells for nerve injury in the central nervous system].

    Science.gov (United States)

    Kinoshita, Hiroyuki

    2013-03-01

    Induced pluripotent stem cells (iPSCs), with both pluripotency and replication competence similar to embryonic stem cells (ESCs), have been developed from mouse fibroblasts in 2006 by Yamanaka et al. iPSCs are unique in employing somatic cells for their production, and can avoid ethical issues existing in ESCs. It is clear that progress in technology to produce iPSCs is one of the most crucial achievements of medicine in this century. Technology with the new pluripotent cells will offer many advantages in the field of regeneration medicine supplying new tissues to the injured organ and/or development of methodology to uncover many genetic diseases. On the other hand, we have to await adequate progress in issues regarding iPSCs, including enhanced efficiency to obtain iPSCs, the technology to produce organs from the cells, avoidance of tumorigenesis and decrease in immunity in response to iPSCs.

  19. Design and fabrication of a nanofibrous polycaprolactone tubular nerve guide for peripheral nerve tissue engineering using a two-pole electrospinning system.

    Science.gov (United States)

    Panahi-Joo, Y; Karkhaneh, A; Nourinia, A; Abd-Emami, B; Negahdari, B; Renaud, P; Bonakdar, S

    2016-04-12

    Nerve guidance conduits are considered to be the new generation of scaffolds designed for nerve disorders. A tubular construct with a highly aligned fibrous structure, mimicking the endoneurium layer surrounding inner axons of a nerve fascicle, is a suitable candidate for a nerve guide. In this paper a new approach for the fabrication of 3D tubular nerve guides is introduced using simulation of a two-pole electrospinning system and describing its mechanism. The structure of this scaffold is then optimized using the Taguchi statistical method and after morphological studies by scanning electron microscopy, the crystallinity, tensile strength and protein adsorption of these highly aligned fibres are investigated, comparing them with semi-aligned and random fibres produced via conventional mandrel electrospinning. Cell attachment, proliferation and migration of PC12 neuronal like cells are studied on highly aligned, semi aligned and random structures, and morphological change and elongation are observed in PC12 cells. The results of these studies suggest that conduits fabricated using two-pole electrospinning are a suitable and promising scaffold for peripheral and even spinal nerve regeneration. This nerve guide has a great potential for further advanced modifications and regeneration in higher levels.

  20. Origin of Matrix-Producing Cells That Contribute to Aortic Fibrosis in Hypertension.

    Science.gov (United States)

    Wu, Jing; Montaniel, Kim Ramil C; Saleh, Mohamed A; Xiao, Liang; Chen, Wei; Owens, Gary K; Humphrey, Jay D; Majesky, Mark W; Paik, David T; Hatzopoulos, Antonis K; Madhur, Meena S; Harrison, David G

    2016-02-01

    Various hypertensive stimuli lead to exuberant adventitial collagen deposition in large arteries, exacerbating blood pressure elevation and end-organ damage. Collagen production is generally attributed to resident fibroblasts; however, other cells, including resident and bone marrow-derived stem cell antigen positive (Sca-1(+)) cells and endothelial and vascular smooth muscle cells, can produce collagen and contribute to vascular stiffening. Using flow cytometry and immunofluorescence, we found that adventitial Sca-1(+) progenitor cells begin to produce collagen and acquire a fibroblast-like phenotype in hypertension. We also found that bone marrow-derived cells represent more than half of the matrix-producing cells in hypertension, and that one-third of these are Sca-1(+). Cell sorting and lineage-tracing studies showed that cells of endothelial origin contribute to no more than one fourth of adventitial collagen I(+) cells, whereas those of vascular smooth muscle lineage do not contribute. Our findings indicate that Sca-1(+) progenitor cells and bone marrow-derived infiltrating fibrocytes are major sources of arterial fibrosis in hypertension. Endothelial to mesenchymal transition likely also contributes, albeit to a lesser extent and pre-existing resident fibroblasts represent a minority of aortic collagen-producing cells in hypertension. This study shows that vascular stiffening represents a complex process involving recruitment and transformation of multiple cells types that ultimately elaborate adventitial extracellular matrix.

  1. Enhancement of insulin-producing cell differentiation from embryonic stem cells using pax4-nucleofection method

    Institute of Scientific and Technical Information of China (English)

    Han-Tso Lin; Hung-Hai Ku; Chung-Lan Kao; Kun-Hsiung Lee; Yuh-Lih Chang; Shih-Hwa Chiou; Fu-Ting Tsai; Tung-Hu Tsai; Dey-Chyi Sheu; Larry LT Ho

    2007-01-01

    AIM: To enhance the differentiation of insulin producing cell (IPC) ability from embryonic stem (ES) cells in vitro.METHODS: Four-day embryoid body (EB)-formatted ES cells were dissociated as single cells for the followed plasmid DNA delivery. The use of Nucleofector- Electroporator (Amaxa biosystems, Germany) in combination with medium-contained G418 provided a high efficiency of gene delivery for advanced selection. Neucleofected cells were plated on the top of fibronectin coated Petri dishes. Addition of Ly294002 and raised the glucose in medium at 24 h before examination.The differentiation status of these cells was monitored by semi-quantitative PCR (SQ-PCR) detection of the expression of relative genes, such as oct-4, sox-17, foxa2, mixl1, pdx-1, insulin 1, glucagons and somatostatin. The percentage of IPC population on d 18 of the experiment was investigated by immunohistochemistry (IHC), and the content/secretion of insulin was estimated by ELISA assay. The mice with severe combined immunodeficiency disease (SCID) pretreated with streptozotocin (STZ) were used to eliminate plasma glucose restoration after pax4+ ES implantation.RESULTS: A high efficiency of gene delivery was demonstrated when neucleofection was used in the present study; approximately 70% cells showed DsRed expression 2 d after neucleofection. By selection of medium-contained G418, the percentage of DsRed expressing cells kept high till the end of study. The pancreatic differentiation seemed to be accelerated by pax4 nucleofection. When compared to the group of cells with mock control, foxa2, mixl1, pdx1, higher insulin and somatostatin levels were detected by SQ-PCR 4 d after nucleofection in the group of pax4 expressing plasmid delivery. Approximately 55% of neucleofected cells showed insulin expression 18 d after neucleofection, and only 18% of cells showed insulin expression in mock control. The disturbance was shown by nucleofected pax4 RNAi vector; only 8% of cells expressed insulin 18

  2. Neurotrophic Effect of Adipose Tissue-Derived Stem Cells on Erectile Function Recovery by Pigment Epithelium-Derived Factor Secretion in a Rat Model of Cavernous Nerve Injury

    Directory of Open Access Journals (Sweden)

    Xin Chen

    2016-01-01

    Full Text Available The paracrine effect is the major mechanism of stem cell therapy. However, the details of the effect’s mechanism remain unknown. The aim of this study is to investigate whether adipose tissue-derived stem cells (ADSCs can ameliorate cavernous nerve injury-induced erectile dysfunction (CNIED rats and to determine its mechanism. Twenty-eight days after intracavernous injection of 5-ethynyl-2-deoxyuridine- (EdU- labeled ADSCs, the erectile function of all the rats was evaluated by intracavernosal pressure (ICP. The ADSCs steadily secreted detectable pigment epithelium-derived factor (PEDF in vitro. The expression of PEDF increased in the penis of the bilateral cavernous nerve injury (BCNI group for 14 days and then gradually decreased. On day 28 after the intracavernous injection, the ADSCs group exhibited a significantly increased ICP compared with the phosphate buffered saline- (PBS- treated group. Moreover, the neuronal nitric oxide synthase (nNOS and S100 expression in penile dorsal nerves and the smooth muscle content to collagen ratio in penile tissues significantly increased. Furthermore, elevated PEDF, p-Akt, and p-eNOS were identified in the ADSCs group. This study demonstrated that intracavernous injection of ADSCs improved erectile function, repaired the nerve, and corrected penile fibrosis. One potential mechanism is the PEDF secretion of ADSCs and subsequent PI3K/Akt pathway activation.

  3. Cutaneous nerve entrapment syndrome

    Institute of Scientific and Technical Information of China (English)

    DongFuhui

    2004-01-01

    The cutaneous nerve entrapment syndrome is named that, the cutaneous nerve's functional disorder caused by some chronic entrapment, moreover appears a series of nerve's feeling obstacle,vegetative nerve function obstacle, nutrition obstacle, even motor function obstacle in various degree.

  4. Nerve biopsy (image)

    Science.gov (United States)

    Nerve biopsy is the removal of a small piece of nerve for examination. Through a small incision, a sample ... is removed and examined under a microscope. Nerve biopsy may be performed to identify nerve degeneration, identify ...

  5. Vagus Nerve Stimulation

    Science.gov (United States)

    Vagus nerve stimulation Overview By Mayo Clinic Staff Vagus nerve stimulation is a procedure that involves implantation of a device that stimulates the vagus nerve with electrical impulses. There's one vagus nerve on ...

  6. Ulnar nerve dysfunction

    Science.gov (United States)

    Neuropathy - ulnar nerve; Ulnar nerve palsy; Mononeuropathy; Cubital tunnel syndrome ... neuropathy occurs when there is damage to the ulnar nerve. This ... syndrome may result. When damage destroys the nerve covering ( ...

  7. Effect of Aβ1-40 on membrane permeability and intracellular free Ca2+ of nerve cells

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    The effects of soluble and fibrillar Aβ1-40 on membrane permeability and intracellular free Ca2+ of nerve cells were investigated by the laser confocal microscopy. Results indicate that: i) Effects of soluble and fibrillar Aβ1-40 on cell membrane permeability are both concentration-de- pendent. Soluble Aβ1-40 increases membrane permeability only at concentration of 3 ?mol/L, while the toxic effect of fibrillar Aβ1-40 is much stronger, its evident effect begins from 1 ?mol/L. When its concentration rose to 3 ?mol/L, not only the membrane permeability increased, but also the nuclear membrane broke seriously. ii) Both soluble and fibrillar Aβ1-40 at high concentrations increased the intracellular free Ca2+, and the increased amplitudes are concentration-dependent. However, the fibrillar one induces the increase of intracellular Ca2+ much quicker and synchronously. These results indicate that some correlation exists between the neurotoxicity of high concentration soluble and fibrillar Aβ1-40 and the change of physico-chemical properties and intracellular Ca ion imbalance.

  8. Zero valent zinc nanoparticles promote neuroglial cell proliferation: A biodegradable and conductive filler candidate for nerve regeneration.

    Science.gov (United States)

    Aydemir Sezer, Umran; Ozturk, Kevser; Aru, Basak; Yanıkkaya Demirel, Gulderen; Sezer, Serdar; Bozkurt, Mehmet Recep

    2017-01-01

    Regeneration of nerve, which has limited ability to undergo self-healing, is one of the most challenging areas in the field of tissue engineering. Regarding materials used in neuroregeneration, there is a recent trend toward electrically conductive materials. It has been emphasized that the capacity of conductive materials to regenerate such tissue having limited self-healing ability improves their clinical utility. However, there have been concerns about the safety of materials or fillers used for conductance due to their lack of degradability. Here, we attempt to use poly(Ɛ-caprolactone) (PCL) matrix consisting of varying proportions of zero valent zinc nanoparticles (Zn NPs) via electrospinning. These conductive, biodegradable, and bioactive materials efficiently promoted neuroglial cell proliferation depending on the amount of Zn NPs present in the PCL matrix. Chemical characterizations indicated that the incorporated Zn NPs do not interact with the PCL matrix chemically and that the Zn NPs improved the tensile properties of the PCL matrix. All composites exhibited linear conductivity under in vitro conditions. In vitro cell culture studies were performed to determine the cytotoxicity and proliferative efficiency of materials containing different proportions of Zn NPs. The results were obtained to explore new conductive fillers that can promote tissue regeneration.

  9. [The interaction between nerve cells and carbon nanotube networks made by CVD process investigation].

    Science.gov (United States)

    Bobrinetskiĭ, I I; Seleznev, A S; Gaĭduchenko, I A; Fedorov, G E; Domantovskiĭ, A G; Presniakov, M Iu; Podcherniaeva, R Ia; Mikhaĭlova, G R; Suetina, I A

    2013-01-01

    In this research we investigate neuroblastoma cells cultivated on single-walled carbon nanotubes networks made by CVD method on silicon substrates. The complex analysis of grown cells made by atomic force, electron microscopy and Raman spectroscopy was carried out and the effect of nanotube growth process on proliferation factor was investigated. It is shown that despite of a weak decrease in proliferation, cell morphology remains unchanged and no physical or chemical interaction between carbon nanotubes and cells is observed. The results of the research can be used to investigate the interaction between conductive nanomaterials and cells for the development of neural replacement implants. Also they can be useful in bio-electronic interface investigation of signal propagation in neurons.

  10. Mitochondrial energy metabolism changes during aging-mouse cranial nerve cells treated with various doses and forms of Fructus schizandrae

    Institute of Scientific and Technical Information of China (English)

    Hongyan Guo; Jinhe Li

    2008-01-01

    BACKGROUND: During the cellular aging process, the number of mitochondria, generation of adenosine triphosphate (ATP), activity of respiratory chain enzyme complex 1 and 4, and oxidation decrease. OBJECTIVE: To observe the effects of aqueous and spirituous extract, as well as polysaccharides from Fructus schizandrae (Magnolia Vine) on energy metabolism and mitochondrial anti-oxidation in cranial nerve cells of a D-gal-induccd aging mouse model.DESIGN, TIME AND SETTING: A randomized, controlled, animal study. The experiment was conducted at the Department of Biochemistry, Qiqihar Medical College between March and July 2006.MATERIALS: Fifty healthy, Kunming mice of both sexes, aged 2 3 months old and weighing 18-22 g, were used for the present study. Fructus schizandrae was purchased from the Medical College of Jiamusi University. Aqueous extracts, spirituous extracts, and polysaccharides from Fructus schizandrae were prepared. D-galactose (D-gal) is a product of the Second Reagent Factory, Shanghai City, China. Mn-superoxide dismutase (Mn-SOD) kit, malonaldehyde (MDA) kit, protein quantification kit, and inorganic phosphorus testing kit were purchased from Jian Cheng Bioeng. Co., China.METHODS: Fifty mice were randomly divided into five groups, with 10 mice in each group: young control, aging model, aqueous Fructus schizandrae extract, spirituous Fructus schizandrae extract, and Fructus schizandrae polysaccharides. Over a course of 30 days, mice in aging model, aqueous Fructus schizandrae extract, spirituous Fructus schizandrae extract, and Fructus schizandrae polysaccharides groups were injected subcutaneously with D-gal (100 mg/kg) into the nape of the neck daily, and administered intragastrically with an equal volume of sterile, warm water (aging model), aqueous Fructus schizandrae extract (2 g/kg), spirituous Fructus schizandrae extract (2 g/kg), or Fructus schizandrae polysaccharides (0.2 g/kg), respectively. Mice in the young control group were injected into

  11. Atypical and classical memory B cells produce Plasmodium falciparum neutralizing antibodies

    DEFF Research Database (Denmark)

    Muellenbeck, Matthias F; Ueberheide, Beatrix; Amulic, Borko

    2013-01-01

    . We show at the single cell level that natural Pf infection induces the development of classical memory B cells (CM) and atypical memory B cells (AtM) that produce broadly neutralizing antibodies against blood stage Pf parasites. CM and AtM contribute to anti-Pf serum IgG production, but only AtM show...

  12. Safety profile of bone marrow mononuclear stem cells in the rehabilitation of patients with posttraumatic facial nerve paralysis-a novel modality (phase one trial).

    Science.gov (United States)

    Aggarwal, Sushil Kumar; Gupta, Ashok Kumar; Modi, Manish; Gupta, Rijuneeta; Marwaha, Neelam

    2012-08-01

    Objectives The objectives of this study were to study the safety profile and role of mononuclear stem cells in the rehabilitation of posttraumatic facial nerve paralysis not improving with conventional treatment. Study Design This is a prospective nonrandomized controlled trial. Study Setting This study is conducted at Postgraduate Institute of Medical Education and Research (PGIMER), Chandigarh between July, 2007 and December, 2008. Patients We included eight patients of either sex aged between 18 and 60 years of posttraumatic facial nerve paralysis not improving with conventional treatment presented to PGIMER, Chandigarh between July 2007 and December 2008. Methods All patients underwent preoperative electroneuronography (ENoG), clinical photography, and high-resolution computed tomography (HRCT) temporal bone. All patients then underwent facial nerve decompression and stem cell implantation. Stem cells processing was done in well-equipped bone marrow laboratory. Postoperatively, all patients underwent repeat ENoG and clinical photography at 3 and 6 months to assess for objective and clinical improvement. Clinical improvement was graded according to modified House-Brackmann grading system. Intervention Done All patients of posttraumatic facial nerve paralysis who were not improving with conventional surgical treatment were subjected to facial nerve decompression and stem cell implantation. Main Outcome Measures All patients who were subjected to stem cell implantation were followed up for 6 months to assess for any adverse effects of stem cell therapy on human beings; no adverse effects were seen in any of our patients after more than 6 months of follow-up. Results Majority of the patients were male, with motor vehicle accidents as the most common cause of injury in our series. Majority had longitudinal fractures on HRCT temporal bone. The significant improvement in ENoG amplitude was seen between preoperative and postoperative amplitudes on

  13. Interleukin 17-Producing γδT Cells Increased in Patients with Active Pulmonary Tuberculosis

    Institute of Scientific and Technical Information of China (English)

    Meiyu Peng; Zhaohua Wang; Chunyan Yao; Lina Jiang; Qili Jin; Jing Wang; Baiqing Li

    2008-01-01

    Although it has been known that y8 T cells may play an important role in the immune response to infection of Mycobacterium tuberculosis (M. tb), the mechanisms by which the T8 T cells participate in the innate and/or acquired immunity to tuberculosis (TB) have not been full elucidated. In the present study, 27 patients with active pulmonary TB and 16 healthy donors (HD) were performed. We found that proportion of IL-17-producing cells among lymphocyte was similar between TB patients and HD, whereas the proportions of γδ T cells in IL-17- producing cells (59.2%) and IL-17-producing cells in γδ T cells (19.4%) in peripheral blood were markedly increased in TB patients when compared to those in HD (43.9% and 7.7%, respectively). In addition, the proportions of IFN-γ-producing γδ T cells in TB patients were obviously lower than that in HD. Upon re-stimulated with M. tb heat-treated antigen (M. tb-HAg) in vitro, fewer IL-17-producing γδ T cells were generated from HD and TB patients, whereas IFN-γ-producing γδ T cells were increased in TB patients compared to that in HD. Our findings in TB patients and healthy human were consistent with other murine investigation that the IL-17- producing γδ T cells were main source of IL-17 in mouse model of BCG infection, suggesting that γδ T cells might be involved in the formation of tubercular granuloma in pulmonary TB patients, but need further identification. Cellular & Molecular Immunology. 2008;5(3):203-208.

  14. Endocrine cells and nerve ganglia of the small intestine of the Opossum Didelphis aurita Wied-Neuwied, 1826 (Mammalia: Didelphidae

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    Gláucia M. Freitas-Ribeiro

    2012-09-01

    Full Text Available The nervous and endocrine systems jointly control intestinal movements, secretions of their glands and also participate of the processes of nutrient digestion and absorption. Therefore, the central objective of this study was to verify the existence of a possible relationship between the number of nervous cells and ganglia of the submucosal and myenteric plexuses and the number of endocrine cells in the small intestine of adult D. aurita. The utilized staining techniques were Grimelius, modified Masson-Fontana, direct immunoperoxidase and H-E. Argyrophillic, argentaffin and insulin immunoreactive endocrine cells do not numerically vary between the initial, mid and final regions of the duodenum, jejunum and ileum (P>0.05, except for argyrophillic cells in the jejunum (P>0.05. No numerical relationship has yet been verified between the number of nerve ganglia and endocrine cells, and also between nervous and endocrine cells. We recommended the use of new immunohistochemical techniques to confirm the numerical correlation between the nervous and endocrine systems in the small intestine. The morphology and distribution of endocrine cells and the nerve ganglia studied were similar to those encountered in eutherian mammals.Os sistemas nervoso e endócrino controlam integra-damente os movimentos intestinais, a secreção de suas glândulas e também participam dos processos de digestão e absorção de nutrientes. Portanto, o objetivo central deste estudo foi verificar a existência de uma possível relação entre o número de células nervosas e gânglios dos plexos submucosos e mioentéricos e o número de células endócrinas no intestino delgado de adultos de D. aurita. As técnicas de coloração utilizadas foram Grimelius, Masson-Fontana modificada, imunoperoxidase direta e H-E. As células endócrinas argirófilas, argentafins e imunorreativas à insulina não variaram numericamente entre as regiões inicial, média e final do duodeno, jejuno e

  15. Regeneração de nervos periféricos: terapia celular e fatores neurotróficos Peripheral nerve regeneration: cell therapy and neurotrophic factors

    Directory of Open Access Journals (Sweden)

    Alessandra Deise Sebben

    2011-01-01

    surgical intervention is rare. Many surgical techniques can be used for nerve repair. Among these, the tubulization technique can be highlighted: this allows regenerative factors to be introduced into the chamber. Cell therapy and tissue engineering have arisen as an alternative for stimulating and aiding peripheral nerve regeneration. Therefore, the aim of this review was to provide a survey and analysis on the results from experimental and clinical studies that used cell therapy and tissue engineering as tools for optimizing the regeneration process. The articles used came from the LILACS, Medline and SciELO scientific databases. Articles on the use of stem cells, Schwann cells, growth factors, collagen, laminin and platelet-rich plasma for peripheral nerve repair were summarized over the course of the review. Based on these studies, it could be concluded that the use of stem cells derived from different sources presents promising results relating to nerve regeneration, because these cells have a capacity for neuronal differentiation, thus demonstrating effective functional results. The use of tubes containing bioactive elements with controlled release also optimizes the nerve repair, thus promoting greater myelination and axonal growth of peripheral nerves. Another promising treatment is the use of platelet-rich plasma, which not only releases growth factors that are important in nerve repair, but also serves as a carrier for exogenous factors, thereby stimulating the proliferation of specific cells for peripheral nerve repair.

  16. Magnetically levitated nano-robots: an application to visualization of nerve cells injuries.

    Science.gov (United States)

    Lou, Mingji; Jonckheere, Edmond

    2007-01-01

    This paper proposes a swarm of magnetically levitated nano-robots with high sensitivity nano-sensors as a mean to detect chemical sources, specifically the chemical signals released by injured nervous cells. In the aftermath of the process, further observation by these nano-robots would be used to monitor the healing process and assess the amount of regeneration, if any, or even the repair, of the injured nervous cells.

  17. A Stem Cell-Seeded Nanofibrous Scaffold for Auditory Nerve Replacement

    Science.gov (United States)

    2015-10-01

    Nanofibrous Substrate, Neural Hearing Loss ESCs - embryonic stem cells EBs - embryoid bodies SMADi - SMAD-inhibitors RA - retinoic acid Pur... embryoid bodies (EBs) are generated by spin-aggregation in a micropatterned plate and pushed toward a neuroectodermal fate using SMAD-inhibitors (SMADi...using a Pax2-reporter cell line. At day 0, mouse ESCs are aggregated into embryoid bodies using round-bottom 96-well plates. On day 1, growth

  18. Delayed peripheral nerve repair: methods, including surgical ′cross-bridging′ to promote nerve regeneration

    Directory of Open Access Journals (Sweden)

    Tessa Gordon

    2015-01-01

    Full Text Available Despite the capacity of Schwann cells to support peripheral nerve regeneration, functional recovery after nerve injuries is frequently poor, especially for proximal injuries that require regenerating axons to grow over long distances to reinnervate distal targets. Nerve transfers, where small fascicles from an adjacent intact nerve are coapted to the nerve stump of a nearby denervated muscle, allow for functional return but at the expense of reduced numbers of innervating nerves. A 1-hour period of 20 Hz electrical nerve stimulation via electrodes proximal to an injury site accelerates axon outgrowth to hasten target reinnervation in rats and humans, even after delayed surgery. A novel strategy of enticing donor axons from an otherwise intact nerve to grow through small nerve grafts (cross-bridges into a denervated nerve stump, promotes improved axon regeneration after delayed nerve repair. The efficacy of this technique has been demonstrated in a rat model and is now in clinical use in patients undergoing cross-face nerve grafting for facial paralysis. In conclusion, brief electrical stimulation, combined with the surgical technique of promoting the regeneration of some donor axons to ′protect′ chronically denervated Schwann cells, improves nerve regeneration and, in turn, functional outcomes in the management of peripheral nerve injuries.

  19. Changes in brain-derived neurotrophic factor expression after transplanting microencapsulated sciatic nerve cells of rabbits into injured spinal cord of rats

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    BACKGROUND: Changes of brain-derived neurotrophic factor (BDNF) expression reflect function of nerve cells; meanwhile, they play a significant role in researching interventions on plerosis of nerve injury.OBJECTIVE: To observe and compare the effects on changes of BDNF expression in rats with spinal cord injury between microencapsulated sciatic nerve cells of rabbits and only transplanting sciatic nerve cells of rabbits.DESIGN: Randomized controlled animal study.SETTING: Medical School of Jiujiang College.MATERIALS: The experiment was carried out in the Medical Science Researching Center, Jiujiang College from May 2004 to May 2006. A total of 90 healthy adult SD rats, weighing 250 - 300 g, of either gender; and 10 rabbits, weighing 2.0 - 2.5 kg, of either gender, were provided by Jiangxi Experimental Animal Center.METHODS: Sciatic nerve tissue of rabbits was separated to make cell suspension. After centrifugation,suspension was mixed with 15 g/L alginate saline solution and ejaculated to 20 mmol/L barium chloride saline solution by double-cavity ejaculator. The obtained cell microcapsules were suspended in saline. Rats were randomly divided into microencapsulated group, only suspension group, and only injured group with 30 animals in each group. After anesthesia, T10 spinous process and vertebra lamina of rats in the former two groups were exposed. Spinal cord tissue in 2-mm length was removed from rats by spinal cord right hemi-section. The gelatin sponges with the size of 2 mm × 2 mm × 2 mm were grafted as filing cage,and absorbed 10 μμ L microencapsulated sciatic nerve cells of rabbit in the microencapsulated group and 10 μ L sciatic nerve cells of rabbits in the only suspension group; respectively. No graft was placed in the only injured group.MAIN OUTCOME MEASURES: On the 1st, 3rd, 7th, 14th and 28th days after operation,immunohistochemistry (SABC technique) was used to detect distribution and amount of positive-reactive neurons in BDNF of spinal cord

  20. Bortezomib Treatment Produces Nocifensive Behavior and Changes in the Expression of TRPV1, CGRP, and Substance P in the Rat DRG, Spinal Cord, and Sciatic Nerve

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    M. Quartu

    2014-01-01

    Full Text Available To investigate neurochemical changes associated with bortezomib-induced painful peripheral neuropathy (PN, we examined the effects of a single-dose intravenous administration of bortezomib and a well-established “chronic” schedule in a rat model of bortezomib-induced PN. The TRPV1 channel and sensory neuropeptides CGRP and substance P (SP were studied in L4-L5 dorsal root ganglia (DRGs, spinal cord, and sciatic nerve. Behavioral measures, performed at the end of the chronic bortezomib treatment, confirmed a reduction of mechanical nociceptive threshold, whereas no difference occurred in thermal withdrawal latency. Western blot analysis showed a relative increase of TRPV1 in DRG and spinal cord after both acute and chronic bortezomib administration. Reverse transcriptase-polymerase chain reaction revealed a decrease of TRPV1 and CGRP mRNA relative levels after chronic treatment. Immunohistochemistry showed that in the DRGs, TRPV1-, CGRP-, and SP-immunoreactive neurons were mostly small- and medium-sized and the proportion of TRPV1- and CGRP-labeled neurons increased after treatment. A bortezomib-induced increase in density of TRPV1- and CGRP-immunoreactive innervation in the dorsal horn was also observed. Our findings show that bortezomib-treatment selectively affects subsets of DRG neurons likely involved in the processing of nociceptive stimuli and that neurochemical changes may contribute to development and persistence of pain in bortezomib-induced PN.

  1. Producing recombinant therapeutic glycoproteins with enhanced sialylation using CHO-gmt4 glycosylation mutant cells

    Science.gov (United States)

    Goh, John SY; Liu, Yingwei; Chan, Kah Fai; Wan, Corrine; Teo, Gavin; Zhang, Peiqing; Zhang, Yuanxing; Song, Zhiwei

    2014-01-01

    Recombinant glycoprotein drugs require proper glycosylation for optimal therapeutic efficacy. Glycoprotein therapeutics are rapidly removed from circulation and have reduced efficacy if they are poorly sialylated. Ricinus communis agglutinin-I (RCA-I) was found highly toxic to wild-type CHO-K1 cells and all the mutants that survived RCA-I treatment contained a dysfunctional N-acetylglucosaminyltransferase I (GnT I) gene. These mutants are named CHO-gmt4 cells. Interestingly, upon restoration of GnT I, the sialylation of a model glycoprotein, erythropoietin, produced in CHO-gmt4 cells was shown to be superior to that produced in wild-type CHO-K1 cells. This addendum summarizes the applicability of this cell line, from transient to stable expression of the recombinant protein, and from a lab scale to an industrial scale perfusion bioreactor. In addition, CHO-gmt4 cells can be used to produce glycoproteins with mannose-terminated N-glycans. Recombinant glucocerebrosidase produced by CHO-gmt4 cells will not require glycan remodeling and may be directly used to treat patients with Gaucher disease. CHO-gmt4 cells can also be used to produce other glycoprotein therapeutics which target cells expressing mannose receptors. PMID:24911584

  2. Dual mTORC1/2 inhibition induces anti-proliferative effect in NF1-associated plexiform neurofibroma and malignant peripheral nerve sheath tumor cells

    Science.gov (United States)

    Hivelin, Mikael; Nusbaum, Patrick; Hubas, Arnaud; Laurendeau, Ingrid; Lantieri, Laurent; Wolkenstein, Pierre; Vidaud, Michel; Pasmant, Eric; Chapuis, Nicolas; Parfait, Béatrice

    2016-01-01

    Approximately 30-50% of individuals with Neurofibromatosis type 1 develop benign peripheral nerve sheath tumors, called plexiform neurofibromas (PNFs). PNFs can undergo malignant transformation to highly metastatic malignant peripheral nerve sheath tumors (MPNSTs) in 5-10% of NF1 patients, with poor prognosis. No effective systemic therapy is currently available for unresectable tumors. In tumors, the NF1 gene deficiency leads to Ras hyperactivation causing the subsequent activation of the AKT/mTOR and Raf/MEK/ERK pathways and inducing multiple cellular responses including cell proliferation. In this study, three NF1-null MPNST-derived cell lines (90-8, 88-14 and 96-2), STS26T sporadic MPNST cell line and PNF-derived primary Schwann cells were used to test responses to AZD8055, an ATP-competitive “active-site” mTOR inhibitor. In contrast to rapamycin treatment which only partially affected mTORC1 signaling, AZD8055 induced a strong inhibition of mTORC1 and mTORC2 signaling in MPNST-derived cell lines and PNF-derived Schwann cells. AZD8055 induced full blockade of mTORC1 leading to an efficient decrease of global protein synthesis. A higher cytotoxic effect was observed with AZD8055 compared to rapamycin in the NF1-null MPNST-derived cell lines with IC50 ranging from 70 to 140 nM and antiproliferative effect was confirmed in PNF-derived Schwann cells. Cell migration was impaired by AZD8055 treatment and cell cycle analysis showed a G0/G1 arrest. Combined effects of AZD8055 and PD0325901 MEK inhibitor as well as BRD4 (BromoDomain-containing protein 4) inhibitors showed a synergistic antiproliferative effect. These data suggest that NF1-associated peripheral nerve sheath tumors are an ideal target for AZD8055 as a single molecule or in combined therapies. PMID:26840085

  3. Dual mTORC1/2 inhibition induces anti-proliferative effect in NF1-associated plexiform neurofibroma and malignant peripheral nerve sheath tumor cells.

    Science.gov (United States)

    Varin, Jennifer; Poulain, Laury; Hivelin, Mikael; Nusbaum, Patrick; Hubas, Arnaud; Laurendeau, Ingrid; Lantieri, Laurent; Wolkenstein, Pierre; Vidaud, Michel; Pasmant, Eric; Chapuis, Nicolas; Parfait, Béatrice

    2016-06-14

    Approximately 30-50% of individuals with Neurofibromatosis type 1 develop benign peripheral nerve sheath tumors, called plexiform neurofibromas (PNFs). PNFs can undergo malignant transformation to highly metastatic malignant peripheral nerve sheath tumors (MPNSTs) in 5-10% of NF1 patients, with poor prognosis. No effective systemic therapy is currently available for unresectable tumors. In tumors, the NF1 gene deficiency leads to Ras hyperactivation causing the subsequent activation of the AKT/mTOR and Raf/MEK/ERK pathways and inducing multiple cellular responses including cell proliferation. In this study, three NF1-null MPNST-derived cell lines (90-8, 88-14 and 96-2), STS26T sporadic MPNST cell line and PNF-derived primary Schwann cells were used to test responses to AZD8055, an ATP-competitive "active-site" mTOR inhibitor. In contrast to rapamycin treatment which only partially affected mTORC1 signaling, AZD8055 induced a strong inhibition of mTORC1 and mTORC2 signaling in MPNST-derived cell lines and PNF-derived Schwann cells. AZD8055 induced full blockade of mTORC1 leading to an efficient decrease of global protein synthesis. A higher cytotoxic effect was observed with AZD8055 compared to rapamycin in the NF1-null MPNST-derived cell lines with IC50 ranging from 70 to 140 nM and antiproliferative effect was confirmed in PNF-derived Schwann cells. Cell migration was impaired by AZD8055 treatment and cell cycle analysis showed a G0/G1 arrest. Combined effects of AZD8055 and PD0325901 MEK inhibitor as well as BRD4 (BromoDomain-containing protein 4) inhibitors showed a synergistic antiproliferative effect. These data suggest that NF1-associated peripheral nerve sheath tumors are an ideal target for AZD8055 as a single molecule or in combined therapies.

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

    Directory of Open Access Journals (Sweden)

    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

  5. Spink2 modulates apoptotic susceptibility and is a candidate gene in the Rgcs1 QTL that affects retinal ganglion cell death after optic nerve damage.

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    Joel A Dietz

    Full Text Available The Rgcs1 quantitative trait locus, on mouse chromosome 5, influences susceptibility of retinal ganglion cells to acute damage of the optic nerve. Normally resistant mice (DBA/2J congenic for the susceptible allele from BALB/cByJ mice exhibit susceptibility to ganglion cells, not only in acute optic nerve crush, but also to chronic inherited glaucoma that is characteristic of the DBA/2J strain as they age. SNP mapping of this QTL has narrowed the region of interest to 1 Mb. In this region, a single gene (Spink2 is the most likely candidate for this effect. Spink2 is expressed in retinal ganglion cells and is increased after optic nerve damage. This gene is also polymorphic between resistant and susceptible strains, containing a single conserved amino acid change (threonine to serine and a 220 bp deletion in intron 1 that may quantitatively alter endogenous expression levels between strains. Overexpression of the different variants of Spink2 in D407 tissue culture cells also increases their susceptibility to the apoptosis-inducing agent staurosporine in a manner consistent with the differential susceptibility between the DBA/2J and BALB/cByJ strains.

  6. Development of polydimethylsiloxane substrates with tunable elastic modulus to study cell mechanobiology in muscle and nerve.

    Directory of Open Access Journals (Sweden)

    Rachelle N Palchesko

    Full Text Available Mechanics is an important component in the regulation of cell shape, proliferation, migration and differentiation during normal homeostasis and disease states. Biomaterials that match the elastic modulus of soft tissues have been effective for studying this cell mechanobiology, but improvements are needed in order to investigate a wider range of physicochemical properties in a controlled manner. We hypothesized that polydimethylsiloxane (PDMS blends could be used as the basis of a tunable system where the elastic modulus could be adjusted to match most types of soft tissue. To test this we formulated blends of two commercially available PDMS types, Sylgard 527 and Sylgard 184, which enabled us to fabricate substrates with an elastic modulus anywhere from 5 kPa up to 1.72 MPa. This is a three order-of-magnitude range of tunability, exceeding what is possible with other hydrogel and PDMS systems. Uniquely, the elastic modulus can be controlled independently of other materials properties including surface roughness, surface energy and the ability to functionalize the surface by protein adsorption and microcontact printing. For biological validation, PC12 (neuronal inducible-pheochromocytoma cell line and C2C12 (muscle cell line were used to demonstrate that these PDMS formulations support cell attachment and growth and that these substrates can be used to probe the mechanosensitivity of various cellular processes including neurite extension and muscle differentiation.

  7. Development of polydimethylsiloxane substrates with tunable elastic modulus to study cell mechanobiology in muscle and nerve.

    Science.gov (United States)

    Palchesko, Rachelle N; Zhang, Ling; Sun, Yan; Feinberg, Adam W

    2012-01-01

    Mechanics is an important component in the regulation of cell shape, proliferation, migration and differentiation during normal homeostasis and disease states. Biomaterials that match the elastic modulus of soft tissues have been effective for studying this cell mechanobiology, but improvements are needed in order to investigate a wider range of physicochemical properties in a controlled manner. We hypothesized that polydimethylsiloxane (PDMS) blends could be used as the basis of a tunable system where the elastic modulus could be adjusted to match most types of soft tissue. To test this we formulated blends of two commercially available PDMS types, Sylgard 527 and Sylgard 184, which enabled us to fabricate substrates with an elastic modulus anywhere from 5 kPa up to 1.72 MPa. This is a three order-of-magnitude range of tunability, exceeding what is possible with other hydrogel and PDMS systems. Uniquely, the elastic modulus can be controlled independently of other materials properties including surface roughness, surface energy and the ability to functionalize the surface by protein adsorption and microcontact printing. For biological validation, PC12 (neuronal inducible-pheochromocytoma cell line) and C2C12 (muscle cell line) were used to demonstrate that these PDMS formulations support cell attachment and growth and that these substrates can be used to probe the mechanosensitivity of various cellular processes including neurite extension and muscle differentiation.

  8. Potential genotoxic effects of GSM-1800 exposure on human cutaneous and nerve cells

    Energy Technology Data Exchange (ETDEWEB)

    Sanchez, S.; Poulletier De Gannes, F.; Haro, E.; Ruffie, G.; Lagroye, I.; Billaudel, B.; Veyret, B. [PIOM laboratory, UMR 5501 CNRS, ENSCPB, 33 -Pessac (France)

    2006-07-01

    Introduction The GSM-1800 signal has been in use for several years in Europe and questions raised about its potential biological effects, in view of the fact that, with respect to GSM-900, the increase in the carrier frequency corresponds to a more superficial absorption in the tissues. Consequently, the skin becomes an even more important target for the absorption of the radiofrequency radiation (R.F.R.) emitted by mobile phones. Nevertheless, brain tissues remain a critical target. Cells In order to determine whether R.F.R. at 1800 MHz could behave as a genotoxic agent, skin and brain cells were exposed to a 217-Hz-modulated GSM-1800 signal and assayed using the comet assay: (1) normal human epidermal keratinocytes (N.H.E.K.) and dermal fibroblasts (N.H.D.F.) which are cutaneous cells from epidermis and dermis respectively, and (2) the S.H. -S.Y.5.Y. and C.H.M.E.-5 human cell lines, which are neuroblastoma and micro-glial cells, respectively. Exposure The R.F.R. exposure system that was used in these experiments was manufactured by I.T. I.S. (Zurich, Switzerland). It consists in two shorted waveguides allowing to run exposed and sham conditions at the same time in the same culture incubator, at 37 Celsius degrees, 5% CO{sub 2}. It is controlled by a software, which provides blind conditions until completion of data analysis. The specific absorption rate (S.A.R.) used was 2 W/kg, corresponding to the public exposure limit recommended by I.C.N.I.R.P. and the exposure duration was 48 hours. Comet assay At the end of the exposure, cells were removed from their Petri dish by trypsin/EDTA treatment, counted and 5 x 10{sup 4} cells were used to detect DNA damage including single DNA breaks. Positive controls were performed using hydrogen peroxidase (1%, 1 hour). The genotoxic effects were detected using the alkaline comet assay kit (Trevigen slides) following the supplier procedure. Under these conditions, 6 independent experiments were performed for each cell type (2

  9. A novel IL-10-producing innate lymphoid cells (ILC10) in a contact hypersensitivity mouse model

    Science.gov (United States)

    Kim, Hyuk Soon; Jang, Jong-Hwa; Lee, Min Bum; Jung, In Duk; Park, Yeong-Min; Kim, Young Mi; Choi, Wahn Soo

    2016-01-01

    The immunoregulatory cytokine Interleukin 10 (IL-10) protein is produced by various cells during the course of inflammatory disorders. Mainly, it downregulates pro-inflammatory cytokines, antigen presentation, and helper T cell activation. In this study, we show that the ratio of IL-10-producing cells was significantly increased in lineage negative (i.e., not T, B, or leukocyte cell lineages) cells than in lineage positive cells in lymphoid and peripheral tissues. We further observed that IL-10-producing innate lymphoid cells (ILCs), here called firstly ILC10, were increased in number in oxazolone-induced contact hypersensitivity (CHS) mice. In detail, IL-10-producing lineage negative cells were elevated in the axillary, inguinal lymph node, and ear tissues of CHS mice. Notably, the cells expressed classical ILC marker proteins such as CD45, CD127, and Sca-1. Altogether, our findings suggest for the first time that ILC10s are present in various physiological settings and could be involved in numerous immune responses as regulatory cells. [BMB Reports 2016; 49(5): 293-296] PMID:26949018

  10. An experimental and theoretical approach to the study of the photoacoustic signal produced by cancer cells

    Directory of Open Access Journals (Sweden)

    Rafael Pérez Solano

    2012-03-01

    Full Text Available The distinctive spectral absorption characteristics of cancer cells make photoacoustic techniques useful for detection in vitro and in vivo. Here we report on our evaluation of the photoacoustic signal produced by a series of monolayers of different cell lines in vitro. Only the melanoma cell line HS936 produced a detectable photoacoustic signal in which amplitude was dependent on the number of cells. This finding appears to be related to the amount of melanin available in these cells. Other cell lines (i.e. HL60, SK-Mel-1, T47D, Hela, HT29 and PC12 exhibited values similar to a precursor of melanin (tyrosinase, but failed to produce sufficient melanin to generate a photoacoustic signal that could be distinguished from background noise. To better understand this phenomenon, we determined a formula for the time-domain photoacoustic wave equation for a monolayer of cells in a non-viscous fluid on the thermoelastic regime. The theoretical results showed that the amplitude and profile of the photoacoustic signal generated by a cell monolayer depended upon the number and distribution of the cells and the location of the point of detection. These findings help to provide a better understanding of the factors involved in the generation of a photoacoustic signal produced by different cells in vitro and in vivo.

  11. The role of Ca 2+-related signaling in photodynamic injury of nerve and glial cells

    Science.gov (United States)

    Lobanov, A. V.; Petin, Y. O.; Uzdensky, A. B.

    2007-05-01

    Photodynamic therapy (PDT) inhibited and irreversibly abolished firing, caused necrosis of neurons, necrosis, apoptosis and proliferation of glial cells in the isolated crayfish stretch receptor. The role in these processes of the central components of Ca 2+-mediated signaling pathway: phospholipase C, calmodulin, calmodulin-dependent kinase II, and protein kinase C was studied using their inhibitors: ET-18, fluphenazine, KN-93, or staurosporine, respectively. ET-18 reduced functional inactivation of neurons, necrosis and apoptosis of glial cells. Fluphenazine and KN-93 reduced PDT-induced necrosis of neurons and glial cells. Staurosporine enhanced PDT-induced glial apoptosis. PDTinduced gliosis was prevented by KN-93 and staurosporine. Therefore, phospholipase C participated in neuron inactivation and glial necrosis and apoptosis. Calmodulin and calmodulin-dependent kinase II were involved in PDT-induced necrosis of neurons and glial cells but not in glial apoptosis. Protein kinase C protected glia from apoptosis and participated in PDT-induced gliosis and loss of neuronal activity. These data may be used for modulation of PDT of brain tumors.

  12. Expressed sequence tag analysis of adult human optic nerve for NEIBank: Identification of cell type and tissue markers

    Directory of Open Access Journals (Sweden)

    Peterson Katherine

    2009-09-01

    Full Text Available Abstract Background The optic nerve is a pure white matter central nervous system (CNS tract with an isolated blood supply, and is widely used in physiological studies of white matter response to various insults. We examined the gene expression profile of human optic nerve (ON and, through the NEIBANK online resource, to provide a resource of sequenced verified cDNA clones. An un-normalized cDNA library was constructed from pooled human ON tissues and was used in expressed sequence tag (EST analysis. Location of an abundant oligodendrocyte marker was examined by immunofluorescence. Quantitative real time polymerase chain reaction (qRT-PCR and Western analysis were used to compare levels of expression for key calcium channel protein genes and protein product in primate and rodent ON. Results Our analyses revealed a profile similar in many respects to other white matter related tissues, but significantly different from previously available ON cDNA libraries. The previous libraries were found to include specific markers for other eye tissues, suggesting contamination. Immune/inflammatory markers were abundant in the new ON library. The oligodendrocyte marker QKI was abundant at the EST level. Immunofluorescence revealed that this protein is a useful oligodendrocyte cell-type marker in rodent and primate ONs. L-type calcium channel EST abundance was found to be particularly low. A qRT-PCR-based comparative mammalian species analysis reveals that L-type calcium channel expression levels are significantly lower in primate than in rodent ON, which may help account for the class-specific difference in responsiveness to calcium channel blocking agents. Several known eye disease genes are abundantly expressed in ON. Many genes associated with normal axonal function, mRNAs associated with axonal transport, inflammation and neuroprotection are observed. Conclusion We conclude that the new cDNA library is a faithful representation of human ON and EST data

  13. Distribution of nitric oxide synthase, nerve growth factor receptor and interstitial cells of Cajal in hirschsprung s disease and its significance

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Objective To investigate the distribution of nitric oxide synthase (NOS), nerve growth factor receptor (NGFR) and interstitial cells of Cajal (ICCs) in Hirschsprung's disease (HD). Methods The distribution of NOS, NGFR and ICCs was studied by using NADPH diaphorase histochemistry, immunohistochemistry with a monoclonal antibody to human NGFR and the specific polyclonal antibody against c-kit in 8 normal controls and 10 cases of HD. Results NOS and NGFR were abundantly present in the myenteric plexus and in ...

  14. Evaluation of Retinal Nerve Fiber Layer and Ganglion Cell Complex in Patients with Optic Neuritis or Neuromyelitis Optica Spectrum Disorders Using Optical Coherence Tomography in a Chinese Cohort

    OpenAIRE

    Guohong Tian; Zhenxin Li; Guixian Zhao; Chaoyi Feng; Mengwei Li; Yongheng Huang; Xinghuai Sun

    2015-01-01

    We evaluate a cohort of optic neuritis and neuromyelitis optica (NMO) spectrum disorders patients in a territory hospital in China. The peripapillary retinal nerve fiber layer (RNFL) and macular ganglion cell complex (GCC) were measured using spectral-domain OCT after 6 months of acute onset. The results showed that both the peripapillary RNFL and macular GCC were significantly thinner in all optic neuritis subtypes compared to controls. In addition, the recurrent optic neuritis and NMO group...

  15. Quantitative studies of the gastrin-producing cells of the human antrum. A methodological study

    DEFF Research Database (Denmark)

    Nielsen, H O; Halken, S; Lorentzen, M

    1980-01-01

    The antral gastrin-producing cells (G-cells) have been identified by the indirect immunoperoxidase technique in two antrum preparations removed due to a recurrent duodenal and gastric ulcer. Morphometric principles were applied to the G-cells with determination of their volume density, numerical....... A method for estimating the total G-cell population and the total G-cell volume in the antrum was developed. In the antrum removed due to a gastric ulcer the number of G-cells was 190 x 10(6) and their total volume 176 mm3....

  16. Demonstration of immunochemical identity between the nerve growth factor-inducible large external (NILE) glycoprotein and the cell adhesion molecule L1

    DEFF Research Database (Denmark)

    Bock, E; Richter-Landsberg, C; Faissner, A

    1985-01-01

    The nerve growth factor-inducible large external (NILE) glycoprotein and the neural cell adhesion molecule L1 were shown to be immunochemically identical. Immunoprecipitation with L1 and NILE antibodies of [3H]fucose-labeled material from culture supernatants and detergent extracts of NGF......]methionine-labeled early post-natal cerebellar cell cultures or [3H]fucose-labeled NGF-treated PC12 cells, all immunoreactivity for NILE antibody could be removed by pre-clearing with L1 antibody and vice versa....

  17. Endovascular Method for Transplantation of Insulin-Producing Cells to the Pancreas Parenchyma in Swine

    OpenAIRE

    Lundberg, J.; Stone-Elander, S.; Zhang, X. -M; Korsgren, Olle; Jonsson, S; Holmin, S.

    2014-01-01

    Insulin-producing cells are transplanted by portal vein injection as an alternative to pancreas transplantation in both clinical and preclinical trials. Two of the main limitations of portal vein transplantation are the prompt activation of the innate immunity and concomitant loss of islets and a small but significant risk of portal vein thrombosis. Furthermore, to mimic physiological release, the insulin-producing cells should instead be located in the pancreas. The trans-vessel wall approac...

  18. Nerve-dependent and -independent events in blastema formation during Xenopus froglet limb regeneration.

    Science.gov (United States)

    Suzuki, Makoto; Satoh, Akira; Ide, Hiroyuki; Tamura, Koji

    2005-10-01

    Blastema formation, the initial stage of epimorphic limb regeneration in amphibians, is an essential process to produce regenerates. In our study on nerve dependency of blastema formation, we used forelimb of Xenopus laevis froglets as a system and applied some histological and molecular approaches in order to determine early events during blastema formation. We also investigated the lateral wound healing in comparison to blastema formation in limb regeneration. Our study confirmed at the molecular level that there are nerve-dependent and -independent events during blastema formation after limb amputation, Tbx5 and Prx1, reliable markers of initiation of limb regeneration, that start to be expressed independently of nerve supply, although their expressions cannot be maintained without nerve supply. We also found that cell proliferation activity, cell survival and expression of Fgf8, Fgf10 and Msx1 in the blastema were affected by denervation, suggesting that these events specific for blastema outgrowth are controlled by the nerve supply. Wound healing, which is thought to be categorized into tissue regeneration, shares some nerve-independent events with epimorphic limb regeneration, although the healing process results in simple restoration of wounded tissue. Overall, our results demonstrate that dedifferentiated blastemal cells formed at the initial phase of limb regeneration must enter the nerve-dependent epimorphic phase for further processes, including blastema outgrowth, and that failure of entry results in a simple redifferentiation as tissue regeneration.

  19. A Stem Cell-Seeded Nanofibrous Scaffold for Auditory Nerve Replacement

    Science.gov (United States)

    2014-10-01

    Scientific , Holliston, MA) syringe pump set at dispensing rate of 0.05 ml/hr. The syringe pump was placed on linear stage programmed to move to a...and new proposals on cell seeding nanofiber scaffolds are being developed. There is no scientific overlap with the current award. C...unaligned (C and E) and aligned (D and F) Matrigel ® coated nanofibers for 19 days and visualised using epifluorescence imaging. Neurons grown on aligned

  20. A simple model of radial nerve injury in the rhesus monkey to evaluate peripheral nerve repair

    Institute of Scientific and Technical Information of China (English)

    Dong Wang; Qingtang Zhu; Xijun Huang; Guo Fu; Liqiang Gu; Xiaolin Liu; Honggang Wang; Jun Hu; Jianhua Yi; Xiaofeng Niu

    2014-01-01

    Current research on bone marrow stem cell transplantation and autologous or xenogenic nerve transplantation for peripheral nerve regeneration has mainly focused on the repair of peripher-al nerve defects in rodents. In this study, we established a standardized experimental model of radial nerve defects in primates and evaluated the effect of repair on peripheral nerve injury. We repaired 2.5-cm lesions in the radial nerve of rhesus monkeys by transplantation of autografts, acellular allografts, or acellular allografts seeded with autologous bone marrow stem cells. Five months after surgery, regenerated nerve tissue was assessed for function, electrophysiology, and histomorphometry. Postoperative functional recovery was evaluated by the wrist-extension test. Compared with the simple autografts, the acellular allografts and allografts seeded with bone marrow stem cells facilitated remarkable recovery of the wrist-extension functions in the rhesus monkeys. This functional improvement was coupled with radial nerve distal axon growth, a higher percentage of neuron survival, increased nerve fiber density and diameter, increased myelin sheath thickness, and increased nerve conduction velocities and peak amplitudes of compound motor action potentials. Furthermore, the quality of nerve regeneration in the bone marrow stem cells-laden allografts group was comparable to that achieved with autografts. The wrist-extension test is a simple behavioral method for objective quantification of peripheral nerve regeneration.

  1. Cellular therapies based on stem cells and their insulin-producing surrogates: a 2015 reality check.

    Science.gov (United States)

    Giannoukakis, Nick; Trucco, Massimo

    2015-05-01

    Stem cell technology has recently gained a substantial amount of interest as one method to create a potentially limitless supply of transplantable insulin-producing cells to treat, and possibly cure diabetes mellitus. In this review, we summarize the state-of-the art of stem cell technology and list the potential sources of stem cells that have been shown to be useful as insulin-expressing surrogates. We also discuss the milestones that have been reached and those that remain to be addressed to generate bona fide beta cell-similar, insulin-producing surrogates. The caveats, limitations, and realistic expectations are also considered for current and future technology. In spite of the tremendous technical advances realized in the past decade, especially in the field of reprogramming adult somatic cells to become stem cells, the state-of-the art still relies on lengthy and cumbersome in vitro culture methods that yield cell populations that are not particularly glucose-responsive when transplanted into diabetic hosts. Despite the current impediments toward clinical translation, including the potential for immune rejection, the availability of technology to generate patient-specific reprogrammable stem cells has, and will be critical for, important insights into the genetics, epigenetics, biology, and physiology of insulin-producing cells in normal and pathologic states. This knowledge could accelerate the time to reach the desired breakthrough for safe and efficacious beta cell surrogates.

  2. Promoting Nerve Regeneration in a Neurotmesis Rat Model Using Poly(DL-lactide-ε-caprolactone Membranes and Mesenchymal Stem Cells from the Wharton’s Jelly: In Vitro and In Vivo Analysis

    Directory of Open Access Journals (Sweden)

    T. Pereira

    2014-01-01

    Full Text Available In peripheral nerves MSCs can modulate Wallerian degeneration and the overall regenerative response by acting through paracrine mechanisms directly on regenerating axons or upon the nerve-supporting Schwann cells. In the present study, the effect of human MSCs from Wharton’s jelly (HMSCs, differentiated into neuroglial-like cells associated to poly (DL-lactide-ε-caprolactone membrane, on nerve regeneration, was evaluated in the neurotmesis injury rat sciatic nerve model. Results in vitro showed successful differentiation of HMSCs into neuroglial-like cells, characterized by expression of specific neuroglial markers confirmed by immunocytochemistry and by RT-PCR and qPCR targeting specific genes expressed. In vivo testing evaluated during the healing period of 20 weeks, showed no evident positive effect of HMSCs or neuroglial-like cell enrichment at the sciatic nerve repair site on most of the functional and nerve morphometric predictors of nerve regeneration although the nociception function was almost normal. EPT on the other hand, recovered significantly better after HMSCs enriched membrane employment, to values of residual functional impairment compared to other treated groups. When the neurotmesis injury can be surgically reconstructed with an end-to-end suture or by grafting, the addition of a PLC membrane associated with HMSCs seems to bring significant advantage, especially concerning the motor function recovery.

  3. ProNGF derived from rat sciatic nerves downregulates neurite elongation and axon specification in PC12 cells

    Directory of Open Access Journals (Sweden)

    Anna Sofia Trigos

    2015-09-01

    Full Text Available Several reports have shown that a sciatic nerve conditioned media (CM causes neuronal-like differentiation in PC12 cells. This differentiation is featured by neurite outgrowth, which are exclusively dendrites, without axon or sodium current induction. In previous studies, our group reported that the CM supplemented with a generic inhibitor for tyrosine kinase receptors (k252a enhanced the CM-induced morphological differentiation upregulating neurite outgrowth, axonal formation and sodium current elicitation. Sodium currents were also induced by depletion of endogenous proNGF from the CM (pNGFd-CM. Given that sodium currents, neurite outgrowth and axon specification are important features of neuronal differentiation, in the current manuscript, first we investigated if proNGF was hindering the full PC12 cell neuronal-like differentiation. Second, we studied the effects of exogenous wild type (pNGFwt and mutated (pNGFmut proNGF isoforms over sodium currents and, whether or not their addition to the pNGFd-CM would prevent sodium current elicitation. Third, we investigated if proNGF was exerting its negative regulation through the sortilin receptor, and for this, the proNGF action was blocked with neurotensin (NT, a factor known to compete with proNGF for sortilin. Thereby, here we show that pNGFd-CM enhanced cell differentiation, cell proportion with long neurites, total neurite length, induced axonal formation and sodium current elicitation. Interestingly, treatment of PC12 cells with wild type or mutated proNGF isoforms elicited sodium currents. Supplementing pNGFd-CM with pNGFmut reduced 35% the sodium currents. On the other hand, pNGFd-CM+pNGFwt induced larger sodium currents than pNGFd-CM. Finally, treatments with CM supplemented with NT showed that sortilin was mediating proNGF negative regulation, since its blocking induced similar effects than the pNGFd-CM treatment. Altogether, our results suggest that proNGF within the CM, is one of the

  4. Size of the Optic Nerve Head and Its Relationship with the Thickness of the Macular Ganglion Cell Complex and Peripapillary Retinal Nerve Fiber Layer in Patients with Primary Open Angle Glaucoma

    Directory of Open Access Journals (Sweden)

    Nobuko Enomoto

    2015-01-01

    Full Text Available Purpose. To evaluate the relationships among the optic nerve head (ONH area, macular ganglion cell complex (mGCC thickness, circumpapillary retinal nerve fiber layer (cpRNFL thickness, and visual field defects in patients with primary open angle glaucoma (POAG. Methods. This retrospective study included 90 eyes of 90 patients with POAG. The ONH area, rim area, mGCC thickness, and cpRNFL thickness were measured using optical coherence tomography. Mean deviation (MD was measured using standard automated perimetry. The relationships among clinical factors including age, refraction, the ONH area, the rim area, the mGCC thickness, the cpRNFL thickness, and MD were evaluated using correlation coefficients and multiple regression analyses. Results. The significant correlation of the ONH area with refraction (r=0.362, P<0.001, the mGCC thickness (r=0.225, P=0.033, and the cpRNFL thickness (r=0.253, P=0.016 was found. Multiple regression analysis showed that the ONH area, rim area, and MD were selected as significant contributing factors to explain the mGCC thickness and cpRNFL thickness. No factor was selected to explain MD. Conclusions. The ONH area, in other words, the disc size itself may affect the mGCC thickness and cpRNFL thickness in POAG patients.

  5. Sulindac derivatives inhibit cell growth and induce apoptosis in primary cells from malignant peripheral nerve sheath tumors of NF1-patients

    Directory of Open Access Journals (Sweden)

    Friedrich Reinhard E

    2004-05-01

    Full Text Available Abstract Background Malignant peripheral nerve sheath tumors (MPNSTs are neoplasms leading to death in most cases. Patients with Neurofibromatosis type 1 have an increased risk of developing this malignancy. The metabolites of the inactive prodrug Sulindac, Sulindac Sulfide and Sulindac Sulfone (Exisulind are new chemopreventive agents that show promising results in the treatment of different cancer types. In this study we examined the antineoplastic effect of these compounds on primary cells derived from two MPNSTs of Neurofibromatosis type 1 patients. Results Exisulind and Sulindac Sulfide showed a dramatic time- and dose-dependent growth inhibitory effect with IC50-values of 120 μM and 63 μM, respectively. The decrease in viability of the tested cells correlated with induction of apoptosis. Treatment with 500 μM Exisulind and 125 μM Sulindac Sulfide for a period of 2 days increased the rate of apoptosis 21-27-fold compared to untreated cells. Reduced expression of RAS-GTP and phosphorylated ERK1/2 was detected in treated MPNST cells. Moreover, elevated levels of phosphorylated SAPK/JNK were found after drug treatment, and low activation of cleaved caspase-3 was seen. Conclusions Our results suggest that this class of compounds may be of therapeutic benefit for Neurofibromatosis type 1 patients with MPNST.

  6. Effects of Nerve Growth Factor and Basic Fibroblast Growth Factor Promote Human Dental Pulp Stem Cells to Neural Differentiation.

    Science.gov (United States)

    Zhang, Jinlong; Lian, Min; Cao, Peipei; Bao, Guofeng; Xu, Guanhua; Sun, Yuyu; Wang, Lingling; Chen, Jiajia; Wang, Yi; Feng, Guijuan; Cui, Zhiming

    2017-04-01

    Dental pulp stem cells (DPSCs) were the most widely used seed cells in the field of neural regeneration and bone tissue engineering, due to their easily isolation, lack of ethical controversy, low immunogenicity and low rates of transplantation rejection. The purpose of this study was to investigate the role of basic fibroblast growth factor (bFGF) and nerve growth factor (NGF) on neural differentiation of DPSCs in vitro. DPSCs were cultured in neural differentiation medium containing NGF and bFGF alone or combination for 7 days. Then neural genes and protein markers were analyzed using western blot and RT-PCR. Our study revealed that bFGF and NGF increased neural differentiation of DPSCs synergistically, compared with bFGF and NGF alone. The levels of Nestin, MAP-2, βIII-tubulin and GFAP were the most highest in the DPSCs + bFGF + NGF group. Our results suggested that bFGF and NGF signifiantly up-regulated the levels of Sirt1. After treatment with Sirt1 inhibitor, western blot, RT-PCR and immunofluorescence staining showed that neural genes and protein markers had markedly decreased. Additionally, the ERK and AKT signaling pathway played a key role in the neural differentiation of DPSCs stimulated with bFGF + NGF. These results suggested that manipulation of the ERK and AKT signaling pathway may be associated with the differentiation of bFGF and NGF treated DPSCs. Our date provided theoretical basis for DPSCs to treat neurological diseases and repair neuronal damage.

  7. Methylglyoxal, the foe and friend of glyoxalase and Trx/TrxR systems in HT22 nerve cells.

    Science.gov (United States)

    Dafre, A L; Goldberg, J; Wang, T; Spiegel, D A; Maher, P

    2015-12-01

    Methylglyoxal (MGO) is a major glycating agent that reacts with basic residues of proteins and promotes the formation of advanced glycation end products (AGEs) which are believed to play key roles in a number of pathologies, such as diabetes, Alzheimer's disease, and inflammation. Here, we examined the effects of MGO on immortalized mouse hippocampal HT22 nerve cells. The endpoints analyzed were MGO and thiol status, the glyoxalase system, comprising glyoxalase 1 and 2 (GLO1/2), and the cytosolic and mitochondrial Trx/TrxR systems, as well as nuclear Nrf2 and its target genes. We found that nuclear Nrf2 is induced by MGO treatment in HT22 cells, as corroborated by induction of the Nrf2-controlled target genes and proteins glutamate cysteine ligase and heme oxygenase 1. Nrf2 knockdown prevented MGO-dependent induction of glutamate cysteine ligase and heme oxygenase 1. The cystine/glutamate antiporter, system xc(-), which is also controlled by Nrf2, was also induced. The increased cystine import (system xc(-)) activity and GCL expression promoted GSH synthesis, leading to increased levels of GSH. The data indicate that MGO can act as both a foe and a friend of the glyoxalase and the Trx/TrxR systems. At low concentrations of MGO (0.3mM), GLO2 is strongly induced, but at high MGO (0.75 mM) concentrations, GLO1 is inhibited and GLO2 is downregulated. The cytosolic Trx/TrxR system is impaired by MGO, where Trx is downregulated yet TrxR is induced, but strong MGO-dependent glycation may explain the loss in TrxR activity. We propose that Nrf2 can be the unifying element to explain the observed upregulation of GSH, GCL, HO1, TrxR1, Trx2, TrxR2, and system xc(-) system activity.

  8. IL-35-producing B cells are critical regulators of immunity during autoimmune and infectious diseases.

    Science.gov (United States)

    Shen, Ping; Roch, Toralf; Lampropoulou, Vicky; O'Connor, Richard A; Stervbo, Ulrik; Hilgenberg, Ellen; Ries, Stefanie; Dang, Van Duc; Jaimes, Yarúa; Daridon, Capucine; Li, Rui; Jouneau, Luc; Boudinot, Pierre; Wilantri, Siska; Sakwa, Imme; Miyazaki, Yusei; Leech, Melanie D; McPherson, Rhoanne C; Wirtz, Stefan; Neurath, Markus; Hoehlig, Kai; Meinl, Edgar; Grützkau, Andreas; Grün, Joachim R; Horn, Katharina; Kühl, Anja A; Dörner, Thomas; Bar-Or, Amit; Kaufmann, Stefan H E; Anderton, Stephen M; Fillatreau, Simon

    2014-03-20

    B lymphocytes have critical roles as positive and negative regulators of immunity. Their inhibitory function has been associated primarily with interleukin 10 (IL-10) because B-cell-derived IL-10 can protect against autoimmune disease and increase susceptibility to pathogens. Here we identify IL-35-producing B cells as key players in the negative regulation of immunity. Mice in which only B cells did not express IL-35 lost their ability to recover from the T-cell-mediated demyelinating autoimmune disease experimental autoimmune encephalomyelitis (EAE). In contrast, these mice displayed a markedly improved resistance to infection with the intracellular bacterial pathogen Salmonella enterica serovar Typhimurium as shown by their superior containment of the bacterial growth and their prolonged survival after primary infection, and upon secondary challenge, compared to control mice. The increased immunity found in mice lacking IL-35 production by B cells was associated with a higher activation of macrophages and inflammatory T cells, as well as an increased function of B cells as antigen-presenting cells (APCs). During Salmonella infection, IL-35- and IL-10-producing B cells corresponded to two largely distinct sets of surface-IgM(+)CD138(hi)TACI(+)CXCR4(+)CD1d(int)Tim1(int) plasma cells expressing the transcription factor Blimp1 (also known as Prdm1). During EAE, CD138(+) plasma cells were also the main source of B-cell-derived IL-35 and IL-10. Collectively, our data show the importance of IL-35-producing B cells in regulation of immunity and highlight IL-35 production by B cells as a potential therapeutic target for autoimmune and infectious diseases. This study reveals the central role of activated B cells, particularly plasma cells, and their production of cytokines in the regulation of immune responses in health and disease.

  9. Schwannomatosis of the sciatic nerve

    Energy Technology Data Exchange (ETDEWEB)

    Yamamoto, Tetsuji; Maruyama, Shigeki; Mizuno, Kosaku [Dept. of Orthopaedic Surgery, Kobe University School of Medicine (Japan)

    2001-02-01

    A 52-year-old woman with schwannomatosis in the left sciatic nerve is presented. The patient had no stigmata of neurofibromatosis (NF) type 1 or 2. Cutaneous or spinal schwannomas were not detected. Magnetic resonance (MR) imaging of the sciatic nerve revealed more than 15 tumors along the course of the nerve. Histological examination revealed schwannomas consisting of Antoni A and B areas. Immunohistochemical study showed most cells reacting intensely for S-100 protein. The patient underwent conservative follow-up treatment due to the minimal symptoms. The relationship of the disease with NF-2 and plexiform schwannoma is discussed. (orig.)

  10. Cerebellar stem cells do not produce neurons and astrocytes in adult mouse

    Energy Technology Data Exchange (ETDEWEB)

    Su, Xin; Guan, Wuqiang; Yu, Yong-Chun; Fu, Yinghui, E-mail: fuyh@fudan.edu.cn

    2014-07-18

    Highlights: • No new neurons and astrocytes are generated in adult mouse cerebellum. • Very few mash1{sup +} or nestin{sup +} stem cells exist, and most of them are quiescent. • Cell proliferation rate is diversified among cerebellar regions and decreases over time. - Abstract: Although previous studies implied that cerebellar stem cells exist in some adult mammals, little is known about whether these stem cells can produce new neurons and astrocytes. In this study by bromodeoxyuridine (BrdU) intraperitoneal (i.p.) injection, we found that there are abundant BrdU{sup +} cells in adult mouse cerebellum, and their quantity and density decreases significantly over time. We also found cell proliferation rate is diversified in different cerebellar regions. Among these BrdU{sup +} cells, very few are mash1{sup +} or nestin{sup +} stem cells, and the vast majority of cerebellar stem cells are quiescent. Data obtained by in vivo retrovirus injection indicate that stem cells do not produce neurons and astrocytes in adult mouse cerebellum. Instead, some cells labeled by retrovirus are Iba1{sup +} microglia. These results indicate that very few stem cells exist in adult mouse cerebellum, and none of these stem cells contribute to neurogenesis and astrogenesis under physiological condition.

  11. Interleukin 4-producing CD4+ T cells in the skin of cats with allergic dermatitis.

    Science.gov (United States)

    Roosje, P J; Dean, G A; Willemse, T; Rutten, V P M G; Thepen, T

    2002-03-01

    Lesional skin of cats with allergic dermatitis has a cellular infiltrate and a CD4/CD8 ratio comparable to that in humans with atopic dermatitis. CD4+ helper T cells and in particular cells belonging to the Th2 subset play an important role in disease pathogenesis in humans. We investigated the cytokine pattern of CD4+ T cells in situ, with special emphasis on the putative presence of cells producing interleukin 4 (IL4), in cats with allergic dermatitis. Immunohistochemical procedures were used to determine that CD4+ T cells in lesional and nonlesional skin of cats with allergic dermatitis can produce IL4, as occurs in humans. Lesional and nonlesional skin of cats with allergic dermatitis had significantly more IL4+ T cells (P = 0.001) than did skin of healthy control cats. Double staining indicated that all IL4+ cells were positive for pan-T or CD4 markers. Double labeling for mast cell chymase and IL4 stained primarily different cells. Western blotting demonstrated cross-reactivity between the antibody against human IL4 and a feline recombinant IL4. These results indicate that IL4 is primarily produced by CD4+ T cells and is also present in clinically uninvolved skin, indicating a role in the pathogenesis of allergic dermatitis in cats.

  12. A Modified Method of Insulin Producing Cells’ Generation from Bone Marrow-Derived Mesenchymal Stem Cells

    Directory of Open Access Journals (Sweden)

    Paweł Czubak

    2014-01-01

    Full Text Available Type 1 diabetes mellitus is a result of autoimmune destruction of pancreatic insulin producing β-cells and so far it can be cured only by insulin injection, by pancreas transplantation, or by pancreatic islet cells’ transplantation. The methods are, however, imperfect and have a lot of disadvantages. Therefore new solutions are needed. The best one would be the use of differentiated mesenchymal stem cells (MSCs. In the present study, we investigated the potential of the bone marrow-derived MSCs line for in vitro differentiation into insulin producing cells (IPSs. We applied an 18-day protocol to differentiate MSCs. Differentiating cells formed cell clusters some of which resembled pancreatic islet-like cells. Using dithizone we confirmed the presence of insulin in the cells. What is more, the expression of proinsulin C-peptide in differentiated IPCs was analyzed by flow cytometry. For the first time, we investigated the influence of growth factors’ concentration on IPCs differentiation efficiency. We have found that an increase in the concentration of growth factors up to 60 ng/mL of β-FGF/EGF and 30 ng/mL of activin A/β-cellulin increases the percentage of IPCs. Further increase of growth factors does not show any increase of the percentage of differentiated cells. Our findings suggest that the presented protocol can be adapted for differentiation of insulin producing cells from stem cells.

  13. Neuronal cell death, nerve growth factor and neurotrophic models: 50 years on.

    Science.gov (United States)

    Bennet, M R; Gibson, W G; Lemon, G

    2002-01-10

    Viktor Hamburger has just died at the age of 100. It is 50 years since he and Rita Levi-Montalcini laid the foundations for the study of naturally occurring cell death and of neurotrophic factors in the nervous system. In a period of less than 10 years, from 1949 to 1958, Hamburger and Levi-Montalcini made the following seminal discoveries: that neuron cell death occurs in dorsal root ganglia, sympathetic ganglia and the cervical column of motoneurons; that the predictions arising from this observation, namely that survival is dependent on the supply of a trophic factor, could be substantiated by studying the effects of a sarcoma on the proliferation of ganglionic processes both in vivo and in vitro; and that the proliferation of these processes could be used as an assay system to isolate the factor. This work provides a short review mostly of the early history of this subject in the context of the Hamburger/Levi-Montalcini paradigm. This acts as an introduction to a consideration of models that have been proposed to account for how the different sources of growth factors provide for the survival of neurons during development. It is suggested that what has been called the 'social-control' model provides the most parsimonious quantitative description of the contribution of trophic factors to neuronal survival, a concept for which we are in debt to Viktor Hamburger and Rita Levi-Montalcini.

  14. Virus-like particle of Macrobrachium rosenbergii nodavirus produced in Spodoptera frugiperda (Sf9) cells is distinctive from that produced in Escherichia coli.

    Science.gov (United States)

    Kueh, Chare Li; Yong, Chean Yeah; Masoomi Dezfooli, Seyedehsara; Bhassu, Subha; Tan, Soon Guan; Tan, Wen Siang

    2016-11-14

    Macrobrachium rosenbergii nodavirus (MrNV) is a virus native to giant freshwater prawn. Recombinant MrNV capsid protein has been produced in Escherichia coli, which self-assembled into virus-like particles (VLPs). However, this recombinant protein is unstable, degrading and forming heterogenous VLPs. In this study, MrNV capsid protein was produced in insect Spodoptera frugiperda (Sf9) cells through a baculovirus system. Dynamic light scattering (DLS) and transmission electron microscopy (TEM) revealed that the recombinant protein produced by the insect cells self-assembled into highly stable, homogenous VLPs each of approximately 40 nm in diameter. Enzyme-linked immunosorbent assay (ELISA) showed that the VLPs produced in Sf9 cells were highly antigenic and comparable to those produced in E. coli. In addition, the Sf9 produced VLPs were highly stable across a wide pH range (2-12). Interestingly, the Sf9 produced VLPs contained DNA of approximately 48 kilo base pairs and RNA molecules. This study is the first report on the production and characterization of MrNV VLPs produced in a eukaryotic system. The MrNV VLPs produced in Sf9 cells were about 10 nm bigger and had a uniform morphology compared with the VLPs produced in E. coli. The insect cell production system provides a good source of MrNV VLPs for structural and immunological studies as well as for host-pathogen interaction studies. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 2016.

  15. Effects of Nerve Growth Factor on Proliferation and DNA Synthesis of Cultured Human Fetal Retinal Pigment Epithelium Cells

    Institute of Scientific and Technical Information of China (English)

    Wensheng Li; Jun Wen; Deyong Jiang; Jianguang Ding

    2002-01-01

    Objective: To investigate the effects of nerve growth factor(NGF)on proliferation and DNAthesis of cultured human fetal retinal pigment epithelium (RPE)cells in vitro.Methods: Primary culture and subculture of human fetal retinal pigment epithelium cellswere established in vitro first. Cultured RPE cells were treated with NGF by variousconcentrations 0μg/L, 50μg/L, 100μg/L, 200μg/L and 300μg/L(final concentration)for 48 hs.After 48 hs, cells proliferation was measured with methyl thiazolyl tetrazolium(MTT)assay method and the amount of DNA was determined by the absorbance at 280nm of nucleic acid & protein analysis.Results: The A values of 100 μg/L, 200 μg/L, 300 μg/L NGF was(0. 213 7 ± 0. 23 3),(0. 218 8 ±0. 018 1), (0. 232 2 ±0. 016 4) as compared with(0. 189 7 ±0. 015 2) of Avalue of 0 μg/L NGF respectively, q value was 3.63,4.40, 6. 42 and P value was0. 015, 0. 000, 0. 000(q-test). The DNA concentrations of 100 μg/L, 200 μg/L, 300μg/L and 400 μg/L NGF was (981. 220 4 ± 123.535 7), (1 375. 848 4 ±244. 471 8),(1 658.707 1 ± 176. 938 1), (2 353.086 3 ±609. 906 4) μg/ml as compared with(666. 818 8 ± 141. 330 2) μg/ml of DNA concentration of 0 μg/L NGF respectively, qvalue was 3.63,8.20,11.47,19.46, P value was 0. 024,0. 000,0. 000,0. 000 (q-test).Conclusion: The data suggested that NGF could stimulate the proliferation and DNAsynthesis of cultured of hRPE cells in vitro in a dose-dependent manner.

  16. Overexpression of cyclooxygenase-2 in malignant peripheral nerve sheath tumor and selective cyclooxygenase-2 inhibitor-induced apoptosis by activating caspases in human malignant peripheral nerve sheath tumor cells.

    Directory of Open Access Journals (Sweden)

    Michiyuki Hakozaki

    Full Text Available BACKGROUND: Cyclooxygenase-2 (COX-2 is a key enzyme in the conversion of arachidonic acid to prostanoids, and its activation is associated with carcinogenesis as well as inflammation. The antitumor effect of selective COX-2 inhibitors has been noted in various malignancies. Malignant peripheral nerve sheath tumor (MPNST is a rare and aggressive soft tissue sarcoma for which effective treatments have not yet been established. The purpose of this study was to investigate a potential therapeutic role of COX-2 in MPNST. METHODS: We evaluated the expression of COX-2 in 44 cases of high-grade MPNST using immunohistochemical staining and compared the staining results with the characteristics and outcome of the patients. We also investigated the antitumor effect of etodolac, a selective COX-2 inhibitor, on MPNST cells in vitro using the MPNST cell line, FMS-1. RESULTS: Overexpression of COX-2 (≥50% positive cells was observed in 29 cases (65.9%, was significantly associated with a poor overall survival (P = 0.0495, and was considered an independent risk factor for a poor outcome by the results of both univariate and multivariate analysis. Etodolac induced apoptosis of FMS-1 cells through the activation of caspase-8, -9, and -3. Moreover, several caspase inhibitors significantly inhibited etodolac-induced apoptosis. CONCLUSIONS: Selective COX-2 inhibitors including etodolac had an antitumor effect on MPNST cells, and their use holds promise as a novel therapeutic strategy for patients with MPNST to improve their prognoses.

  17. Thymoproteasomes produce unique peptide motifs for positive selection of CD8(+) T cells.

    Science.gov (United States)

    Sasaki, Katsuhiro; Takada, Kensuke; Ohte, Yuki; Kondo, Hiroyuki; Sorimachi, Hiroyuki; Tanaka, Keiji; Takahama, Yousuke; Murata, Shigeo

    2015-01-01

    Positive selection in the thymus provides low-affinity T-cell receptor (TCR) engagement to support the development of potentially useful self-major histocompatibility complex class I (MHC-I)-restricted T cells. Optimal positive selection of CD8(+) T cells requires cortical thymic epithelial cells that express β5t-containing thymoproteasomes (tCPs). However, how tCPs govern positive selection is unclear. Here we show that the tCPs produce unique cleavage motifs in digested peptides and in MHC-I-associated peptides. Interestingly, MHC-I-associated peptides carrying these tCP-dependent motifs are enriched with low-affinity TCR ligands that efficiently induce the positive selection of functionally competent CD8(+) T cells in antigen-specific TCR-transgenic models. These results suggest that tCPs contribute to the positive selection of CD8(+) T cells by preferentially producing low-affinity TCR ligand peptides.

  18. Detection of PIVKA II produced by human hepatoma cells in nude mice.

    Science.gov (United States)

    Kohda, H; Ono, M; Sekiya, C; Ohta, H; Ohhira, M; Ohhira, M; Yoshida, Y; Ikeda, N; Namiki, M

    1991-03-01

    A novel experimental nude mouse model, which is useful for investigation of the mechanisms of PIVKA II synthesis, was established by inoculation with PIVKA II-producing human hepatoma cells (huH-1). We have found markedly elevated levels of PIVKA II in the plasma of nude mice transplanted with huH-1 cells and increased PIVKA II content in huH-1 tumor tissues. Whereas we have not found detectable level of PIVKA II neither in the plasma nor in tumor tissues of nude mice transplanted different human hepatoma cells (HLF) which is not producing PIVKA II. Histology of the tumor tissues produced by huH-1 cells revealed a thick trabecular pattern with blood spaces.

  19. Visualization and identification of IL-7 producing cells in reporter mice.

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    Renata I Mazzucchelli

    Full Text Available Interleukin-7 (IL-7 is required for lymphocyte development and homeostasis although the actual sites of IL-7 production have never been clearly identified. We produced a bacterial artificial chromosome (BAC transgenic mouse expressing ECFP in the Il7 locus. The construct lacked a signal peptide and ECFP (enhanced cyan fluorescent protein accumulated inside IL-7-producing stromal cells in thoracic thymus, cervical thymus and bone marrow. In thymus, an extensive reticular network of IL-7-containing processes extended from cortical and medullary epithelial cells, closely contacting thymocytes. Central memory CD8 T cells, which require IL-7 and home to bone marrow, physically associated with IL-7-producing cells as we demonstrate by intravital imaging.

  20. Neurotrophic factor changes in the rat thick skin following chronic constriction injury of the sciatic nerve

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    Peleshok Jennifer C

    2012-01-01

    Full Text Available Abstract Background Cutaneous peripheral neuropathies have been associated with changes of the sensory fiber innervation in the dermis and epidermis. These changes are mediated in part by the increase in local expression of trophic factors. Increase in target tissue nerve growth factor has been implicated in the promotion of peptidergic afferent and sympathetic efferent sprouting following nerve injury. The primary source of nerve growth factor is cells found in the target tissue, namely the skin. Recent evidence regarding the release and extracellular maturation of nerve growth factor indicate that it is produced in its precursor form and matured in the extracellular space. It is our hypothesis that the precursor form of nerve growth factor should be detectable in those cell types producing it. To date, limitations in available immunohistochemical tools have restricted efforts in obtaining an accurate distribution of nerve growth factor in the skin of naïve animals and those with neuropathic pain lesions. It is the objective of this study to delineate the distribution of the precursor form of nerve growth factor to those cell types expressing it, as well as to describe its distribution with respect to those nerve fibers responsive to it. Results We observed a decrease in peptidergic fiber innervation at 1 week after the application of a chronic constriction injury (CCI to the sciatic nerve, followed by a recovery, correlating with TrkA protein levels. ProNGF expression in CCI animals was significantly higher than in sham-operated controls from 1-4 weeks post-CCI. ProNGF immunoreactivity was increased in mast cells at 1 week post-CCI and, at later time points, in keratinocytes. P75 expression within the dermis and epidermis was significantly higher in CCI-operated animals than in controls and these changes were localized to neuronal and non-neuronal cell populations using specific markers for each. Conclusions We describe proNGF expression by

  1. Staphylococcus aureus produces membrane-derived vesicles that induce host cell death.

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    Mamata Gurung

    Full Text Available Gram-negative bacteria produce outer membrane vesicles that play a role in the delivery of virulence factors to host cells. However, little is known about the membrane-derived vesicles (MVs produced by gram-positive bacteria. The present study examined the production of MVs from Staphylococcus aureus and investigated the delivery of MVs to host cells and subsequent cytotoxicity. Four S. aureus strains tested, two type strains and two clinical isolates, produced spherical nanovesicles during in vitro culture. MVs were also produced during in vivo infection of a clinical S. aureus isolate in a mouse pneumonia model. Proteomic analysis showed that 143 different proteins were identified in the S. aureus-derived MVs. S. aureus MVs were interacted with the plasma membrane of host cells via a cholesterol-rich membrane microdomain and then delivered their component protein A to host cells within 30 min. Intact S. aureus MVs induced apoptosis of HEp-2 cells in a dose-dependent manner, whereas lysed MVs neither delivered their component into the cytosol of host cells nor induced cytotoxicity. In conclusion, this study is the first report that S. aureus MVs are an important vehicle for delivery of bacterial effector molecules to host cells.

  2. Distribution of glutamic acid decarboxylase messenger RNA-containing nerve cell populations of the male rat brain.

    Science.gov (United States)

    Ferraguti, F; Zoli, M; Aronsson, M; Agnati, L F; Goldstein, M; Filer, D; Fuxe, K

    1990-01-01

    The distribution of glutamic acid decarboxylase (GAD) mRNA was investigated throughout the rat brain by means of in situ hybridization. Hybridization was carried out with a 35S-radiolabeled cRNA probe transcribed from a cDNA from cat occipital cortex and cloned in a SP6-T7 promoter-containing vector. Fixed tissue sections were hybridized with 35S GAD probe (0.6 kb length). Signal was detected by means of film or emulsion autoradiography. The autoradiograms were semiquantitatively evaluated by means of computer-assisted image analysis. The results obtained with this evaluation were correlated with the results of the semiquantitative analysis of GAD immunoreactivity performed by Mugnaini and Oertel. Specific labeling was only observed in neuronal cell bodies, whereas no labeling was found over neuropil, glial and endothelial cells. The highest labeling was found in the bulbus olfactorius (internal plexiform and granular layers) and in the caudal magnocellular nucleus of the hypothalamus. Strong labeling was observed in the Purkinje layer of the cerebellar cortex, the interpeduncular nucleus, the interstitial nucleus of Cajal, the nucleus of Darkschewitsch and the suprachiasmatic nucleus. Intermediate or low levels of GAD mRNA were present in various brain nuclei, where gamma-aminobutyric acid (GABA)-containing cell bodies had been observed with other techniques. Interestingly, a low level of GAD mRNA was found in the caudate-putamen and nucleus accumbens, where the vast majority of nerve cells is known to contain GAD immunoreactivity. Only a poor correlation was found between the present semiquantitative measurements of GAD mRNA content and previous analyses of the number of GAD-immunoreactive cell bodies. The present study demonstrates that there exists a differential regional expression of GAD mRNA. The comparison with cell counts performed by immunocytochemistry suggests that some brain areas, such as caudate-putamen and nucleus accumbens, contain a large number

  3. Effect of electrical convulsions on uridine labeling and activity pattern in nerve cells in mice

    Energy Technology Data Exchange (ETDEWEB)

    Pakkenberg, H.; Pakkenberg, B.; Fog, R.

    1985-07-01

    Male white mice were exposed to electroshock and then injected intravenously with 5-(3H)uridine immediately after the shock. After 5, 30, or 60 min or 6, 12, or 24 h, the mice were killed, microautoradiographs were prepared, and grains were counted in the cortex, hippocampus, and basal ganglia. The results of the grain counts were compared with grain counts in the cortex, hippocampus, and basal ganglia of mice exposed to anoxia for 25 s and then treated in the same manner as the first groups. After electroshock the grain count decreased to 25% of that in control animals in the hippocampus and to 50% in the cortex but was normal in the basal ganglia. The counts returned to normal values within 6 h in the hippocampus, and within 1 h in the cortex. After anoxia, the grain counts were normal in the cortex and hippocampus but increased in the basal ganglia. The distribution of cells with a high or low grain count in vertical and horizontal columns of the cortex in control and convulsion animals was analyzed. There were random variations from column to column in both control and convulsion animals. In some anatomic layers there were significantly different grain counts, indicating differences in functional activity.

  4. Vitamin A Impairs the Reprogramming of Tregs into IL-17-Producing Cells during Intestinal Inflammation

    Science.gov (United States)

    Tejón, Gabriela; Manríquez, Valeria; De Calisto, Jaime; Flores-Santibáñez, Felipe; Hidalgo, Yessia; Crisóstomo, Natalia; Fernández, Dominique; Sauma, Daniela; Mora, J. Rodrigo; Bono, María R.; Rosemblatt, Mario

    2015-01-01

    Maintaining the identity of Foxp3+ regulatory T cells (Tregs) is critical for controlling immune responses in the gut, where an imbalance between Tregs and T effector cells has been linked to inflammatory bowel disease. Accumulating evidence suggests that Tregs can convert into Th17 cells and acquire an inflammatory phenotype. In this study, we used an adoptive transfer model of Ag-specific T cells to study the contribution of different factors to the reprogramming of in vitro-generated Treg cells (iTreg) into IL-17-producing cells in a mouse model of gut inflammation in vivo. Our results show that intestinal inflammation induces the reprogramming of iTreg cells into IL-17-producing cells and that vitamin A restrains reprogramming in the gut. We also demonstrate that the presence of IL-2 during the in vitro generation of iTreg cells confers resistance to Th17 conversion but that IL-2 and retinoic acid (RA) cooperate to maintain Foxp3 expression following stimulation under Th17-polarizing conditions. Additionally, although IL-2 and RA differentially regulate the expression of different Treg cell suppressive markers, Treg cells generated under different polarizing conditions present similar suppressive capacity. PMID:26583087

  5. In vitro pancreas duodenal homeobox-1 enhances the differentiation of pancreatic ductal epithelial cells into insulin-producing cells

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    AIM: To observe whether pancreatic and duodenal homeobox factor-1 enhances the differentiation of pancreatic ductal epithelial cells into insulin-producing cells in vitro.METHODS: Rat pancreatic tissue was submitted to digestion by collegenase, ductal epithelial cells were separated by density gradient centrifugation and then cultured in RPMI1640 medium with 10% fetal bovine serum. After 3-5 passages, the cells were incubated in a six-well plate for 24 h before transfection of recombination plasmid XIHbox8VP16. Lightcycler quantitative real-time RT-PCR was used to detect the expression of PDX-1 and insulin mRNA in pancreatic epithelial cells. The expression of PDX-1 and insulin protein was analyzed by Western blotting. Insulin secretion was detected by radioimmunoassay. Insulinproducing cells were detected by dithizone-staining.RESULTS: XIHbox8 mRNA was expressed in pancreatic ductal epithelial cells. PDX-1 and insulin mRNA as well as PDX-1 and insulin protein were significantly increased in the transfected group. The production and insulin secretion of insulin-producing cells differentiated from pancreatic ductal epithelial cells were higher than those of the untransfected cells in vitro with a significant difference (1.32 ± 0.43 vs 3.48 ± 0.81, P < 0.01 at 5.6 mmol/L; 4.86 ± 1.15 vs 10.25 ± 1.32, P < 0.01 at 16.7 mmol/L).CONCLUSION: PDX-1 can differentiate rat pancreatic ductal epithelial cells into insulin-producing cells in vitro.In vitro PDX-1 transfection is a valuable strategy for increasing the source of insulin-producing cells.

  6. Osteopontin is induced by TGF-β2 and regulates metabolic cell activity in cultured human optic nerve head astrocytes.

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    Carolin Neumann

    Full Text Available The aqueous humor (AH component transforming growth factor (TGF-β2 is strongly correlated to primary open-angle glaucoma (POAG, and was shown to up-regulate glaucoma-associated extracellular matrix (ECM components, members of the ECM degradation system and heat shock proteins (HSP in primary ocular cells. Here we present osteopontin (OPN as a new TGF-β2 responsive factor in cultured human optic nerve head (ONH astrocytes. Activation was initially demonstrated by Oligo GEArray microarray and confirmed by semiquantitative (sq RT-PCR, realtime RT-PCR and western blot. Expressions of most prevalent OPN receptors CD44 and integrin receptor subunits αV, α4, α 5, α6, α9, β1, β3 and β5 by ONH astrocytes were shown by sqRT-PCR and immunofluorescence labeling. TGF-β2 treatment did not affect their expression levels. OPN did not regulate gene expression of described TGF-β2 targets shown by sqRT-PCR. In MTS-assays, OPN had a time- and dose-dependent stimulating effect on the metabolic activity of ONH astrocytes, whereas TGF-β2 significantly reduced metabolism. OPN signaling via CD44 mediated a repressive outcome on metabolic activity, whereas signaling via integrin receptors resulted in a pro-metabolic effect. In summary, our findings characterize OPN as a TGF-β2 responsive factor that is not involved in TGF-β2 mediated ECM and HSP modulation, but affects the metabolic activity of astrocytes. A potential involvement in a protective response to TGF-β2 triggered damage is indicated, but requires further investigation.

  7. Upregulated gene expression of local brain-derived neurotrophic factor and nerve growth factor after intracisternal administration of marrow stromal cells in rats with traumatic brain injury

    Institute of Scientific and Technical Information of China (English)

    胡德志; 周良辅; 朱剑虹; 毛颖; 吴雪海

    2005-01-01

    Objective: To examine the effects of rat marrow stromal cells (rMSCs) on gene expression of local brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) after injection of rMSCs into Cistern Magnum of adult rats subjected to traumatic brain injury(TBI).Results: Group cell transplantation had higher BDNF and NGF gene expressions than Group saline control during a period of less than 3 weeks (P<0.05).Conclusions: rMSCs transplantation via Cistern Magnum in rats subjected to traumatic brain injury can enhance expressions of local brain NGF and BDNF to a certain extent.

  8. Retinoic acid primes human dendritic cells to induce gut-homing, IL-10-producing regulatory T cells

    NARCIS (Netherlands)

    Bakdash, G.; Vogelpoel, L.T.; Capel, T.M. van; Kapsenberg, M.L.; Jong, E.C. de

    2015-01-01

    The vitamin A metabolite all-trans retinoic acid (RA) is an important determinant of intestinal immunity. RA primes dendritic cells (DCs) to express CD103 and produce RA themselves, which induces the gut-homing receptors alpha4beta7 and CCR9 on T cells and amplifies transforming growth factor (TGF)-

  9. Liver dendritic cells present bacterial antigens and produce cytokines upon Salmonella encounter.

    Science.gov (United States)

    Johansson, Cecilia; Wick, Mary Jo

    2004-02-15

    The capacity of murine liver dendritic cells (DC) to present bacterial Ags and produce cytokines after encounter with Salmonella was studied. Freshly isolated, nonparenchymal liver CD11c(+) cells had heterogeneous expression of MHC class II and CD11b and a low level of CD40 and CD86 expression. Characterization of liver DC subsets revealed that CD8alpha(-)CD4(-) double negative cells constituted the majority of liver CD11c(+) ( approximately 85%) with few cells expressing CD8alpha or CD4. Flow cytometry analysis of freshly isolated CD11c(+) cells enriched from the liver and cocultured with Salmonella expressing green fluorescent protein (GFP) showed that CD11c(+) MHC class II(high) cells had a greater capacity to internalize Salmonella relative to CD11c(+) MHC class II(low) cells. Moreover, both CD8alpha(-) and CD8alpha(+) liver DC internalized bacteria with similar efficiency after both in vitro and in vivo infection. CD11c(+) cells enriched from the liver could also process Salmonella for peptide presentation on MHC class I and class II to primary, Ag-specific T cells after internalization requiring actin cytoskeletal rearrangements. Flow cytometry analysis of liver CD11c(+) cells infected with Salmonella expressing GFP showed that both CD8alpha(-) and CD8alpha(+) DC produced IL-12p40 and TNF-alpha. The majority of cytokine-positive cells did not contain bacteria (GFP(-)) whereas only a minor fraction of cytokine-positive cells were GFP(+). Furthermore, only approximately 30-50% of liver DC containing bacteria (GFP(+)) produced cytokines. Thus, liver DC can internalize and process Salmonella for peptide presentation to CD4(+) and CD8(+) T cells and elicit proinflammatory cytokine production upon Salmonella encounter, suggesting that DC in the liver may contribute to immunity against hepatotropic bacteria.

  10. From the regulatory functions of B cells to the identification of cytokine-producing plasma cell subsets.

    Science.gov (United States)

    Dang, Van Duc; Hilgenberg, Ellen; Ries, Stefanie; Shen, Ping; Fillatreau, Simon

    2014-06-01

    B lymphocytes have a unique role as antibody-producing cells. Antibodies are key mediators of humoral immunity against infections, and are thought to account for the protection afforded by successful vaccines. B cells can also secrete cytokines and subsequently regulate immune responses mediated by T and innate cells. Remarkably, recent studies identified plasma blasts/plasma cells as the main types of activated B cells producing the cytokines interleukin (IL)-10, IL-35, tumor necrosis factor (TNF)-α, IL-17, and GM-CSF in various contexts in mice. Here, we discuss these observations, which suggest the existence of various subsets of plasma blast/plasma cells distinguishable through their cytokine expression pattern.

  11. Optic Nerve Decompression for Orbitofrontal Fibrous Dysplasia

    OpenAIRE

    Abe, Takumi; Sato, Kaneshige; Otsuka, Takaharu; Kawamura, Noriyoshi; Shimazu, Motohiko; Izumiyama, Hitoshi; Matsumoto, Kiyoshi

    2002-01-01

    Orbitofrontal fibrous dysplasia often involves the bony orbit and the optic canal. Although fibrous dysplasia reportedly produces compression of the optic nerve leading to visual distrubances, optic nerve decompression in patients without clinical signs of optic neuropathy is still controversial. We describe two patients with orbitofrontal fibrous dysplasia without signs of visual disturbance and one patient with McCune-Albright syndrome and progressive visual impairment. Optic nerve decompre...

  12. Fusion of proinsulin-producing bone marrow-derived cells with hepatocytes in diabetes.

    Science.gov (United States)

    Fujimiya, Mineko; Kojima, Hideto; Ichinose, Masumi; Arai, Ryohachi; Kimura, Hiroshi; Kashiwagi, Atsunori; Chan, Lawrence

    2007-03-06

    We previously reported that diabetes in mice is associated with the appearance of proinsulin-producing (Proins-P) cells in the liver. It was unclear, however, whether these Proins-P bone marrow-derived cells (BMDC) merely transit through the liver or undergo fusion with hepatocytes, normally an extremely rare event. In this study, we found that, in diabetes, BMDC in the liver produce not only Proins but also TNF-alpha, suggesting that diabetes reprograms gene expression in BMDC, turning on "inappropriate" genes. Bone marrow transplantation using genetically marked donor and recipient mice showed that fusion occurs between Proins-P BMDC and hepatocytes. Cell fusion is further supported by the presence of the Y chromosome in Proins-P cells in female mice that received male bone marrow transplantation cells. Morphologically, Proins-P fusion cells are albumin-producing hepatocytes that constitute approximately 2.5% of the liver section area 5 months after diabetes induction. An extensive search failed to reveal any fusion cells in nondiabetic mice. Thus, diabetes causes fusion between Proins-P BMDC and hepatocytes in vivo, an observation that has implications for the pathophysiology of diabetes as well as the fundamental biology of heterotypic cell fusion.

  13. Reversal of hyperglycemia in mice by using human expandable insulin-producing cells differentiated from fetal liver progenitor cells

    Science.gov (United States)

    Zalzman, Michal; Gupta, Sanjeev; Giri, Ranjit K.; Berkovich, Irina; Sappal, Baljit S.; Karnieli, Ohad; Zern, Mark A.; Fleischer, Norman; Efrat, Shimon

    2003-06-01

    Beta-cell replacement is considered to be the most promising approach for treatment of type 1 diabetes. Its application on a large scale is hindered by a shortage of cells for transplantation. Activation of insulin expression, storage, and regulated secretion in stem/progenitor cells offers novel ways to overcome this shortage. We explored whether fetal human progenitor liver cells (FH) could be induced to differentiate into insulin-producing cells after expression of the pancreatic duodenal homeobox 1 (Pdx1) gene, which is a key regulator of pancreatic development and insulin expression in beta cells. FH cells possess a considerable replication capacity, and this was further extended by introduction of the gene for the catalytic subunit of human telomerase. Immortalized FH cells expressing Pdx1 activated multiple beta-cell genes, produced and stored considerable amounts of insulin, and released insulin in a regulated manner in response to glucose. When transplanted into hyperglycemic immunodeficient mice, the cells restored and maintained euglycemia for prolonged periods. Quantitation of human C-peptide in the mouse serum confirmed that the glycemia was normalized by the transplanted human cells. This approach offers the potential of a novel source of cells for transplantation into patients with type 1 diabetes.

  14. In vitro cultivation of human fetal pancreatic ductal stem cells and their differentiation into insulin-producing cells

    Institute of Scientific and Technical Information of China (English)

    Zhong-Xiang Yao; Mao-Lin Qin; Jian-Jun Liu; Xing-Shu Chen; De-Shan Zhou

    2004-01-01

    AIM: To isolate, culture and identify the human fetal pancreatic ductal stem cells in vitro, and to observe the potency of these multipotential cells differentiation into insulin-producing cells.METHODS: The human fetal pancreas was digested by 1 g/L collagease type Ⅳ and then 2.5 g/L trypsin was used to isolate the pancreatic ducta stem cells, followed by culture in serum-free, glucose-free DMEM media with some additional chemical substrates in vitro (according to the different Stage). The cells were induced by glucose-free (control),5 mmol/L, 17.8 mmol/L and 25 mmol/L glucose, respectively.The cell types of differentiated cells were identified using immunocytochemical staining.RESULTS: The shape of human fetal pancreatic ductal stem cells culturedin vitro was firstly fusiform in the first 2 wk,and became monolayer and cobblestone pattern after another 3 to 4 wk. After induced and differentiated by the glucose of different concentrations for another 1 to 2 wk,the cells formed the pancreatic islet-like structures. The identification and potency of these cells were then identified by using the pancreatic ductal stem cell marker, cytokeratin-19 (CK-19), pancreatic β cell marker, insulin and pancreatic α cell marker, glucagons with immunocytochemical staining.At the end of the second week, 95.2% of the cells were positive for CK-19 immunoreactivity. Up to 22.7% of the cells induced by glucose were positive for insulin immunoreactivity, and less than 3.8% of the cells were positive for glucagon immunoreactivity in pancreatic isletlike structures. The positive ratio of immunoreactive staining was dependent on the concentration of glucose, and it was observed that the 17.8 mmol/L glucose stimulated effectively to produce insulin- and glucagons-producing cells.CONCLUSION: The human fetal pancreatic ductal stem cells are capable of proliferation in vitro. These cells have multidifferentiation potential and can be induced by glucose and differentiated into insulin-producing

  15. Programmed cell death in barley aleurone cells is not directly stimulated by reactive oxygen species produced in response to gibberellin.

    Science.gov (United States)

    Aoki, Nozomi; Ishibashi, Yushi; Kai, Kyohei; Tomokiyo, Reisa; Yuasa, Takashi; Iwaya-Inoue, Mari

    2014-05-01

    The cereal aleurone layer is a secretory tissue that produces enzymes to hydrolyze the starchy endosperm during germination. We recently demonstrated that reactive oxygen species (ROS), produced in response to gibberellins (GA), promoted GAMyb expression, which induces α-amylase expression in barley aleurone cells. On the other hand, ROS levels increase during programmed cell death (PCD) in barley aleurone cells, and GAMyb is involved in PCD of these cells. In this study, we investigated whether the ROS produced in response to GA regulate PCD directly by using mutants of Slender1 (SLN1), a DELLA protein that negatively regulates GA signaling. The wild-type, the sln1c mutant (which exhibits gibberellin-type signaling even in the absence of GA), and the Sln1d mutant (which is gibberellin-insensitive with respect to α-amylase production) all produced ROS in response to GA, suggesting that ROS production in aleurone cells in response to GA is independent of GA signaling through this DELLA protein. Exogenous GA promoted PCD in the wild-type. PCD in sln1c was induced even without exogenous GA (and so without induction of ROS), whereas PCD in Sln1d was not induced in the presence of exogenous GA, even though the ROS content increased significantly in response to GA. These results suggest that PCD in barley aleurone cells is not directly stimulated by ROS produced in response to GA but is regulated by GA signaling through DELLA protein.

  16. Generation of a Chinese Hamster Ovary Cell Line Producing Recombinant Human Glucocerebrosidase

    Directory of Open Access Journals (Sweden)

    Juliana Branco Novo

    2012-01-01

    Full Text Available Impaired activity of the lysosomal enzyme glucocerebrosidase (GCR results in the inherited metabolic disorder known as Gaucher disease. Current treatment consists of enzyme replacement therapy by administration of exogenous GCR. Although effective, it is exceptionally expensive, and patients worldwide have a limited access to this medicine. In Brazil, the public healthcare system provides the drug free of charge for all Gaucher’s patients, which reaches the order of $ 84 million per year. However, the production of GCR by public institutions in Brazil would reduce significantly the therapy costs. Here, we describe a robust protocol for the generation of a cell line producing recombinant human GCR. The protein was expressed in CHO-DXB11 (dhfr− cells after stable transfection and gene amplification with methotrexate. As expected, glycosylated GCR was detected by immunoblotting assay both as cell-associated (~64 and 59 kDa and secreted (63–69 kDa form. Analysis of subclones allowed the selection of stable CHO cells producing a secreted functional enzyme, with a calculated productivity of 5.14 pg/cell/day for the highest producer. Although being laborious, traditional methods of screening high-producing recombinant cells may represent a valuable alternative to generate expensive biopharmaceuticals in countries with limited resources.

  17. B cells produce less IL-10, IL-6 and TNF-α in myasthenia gravis.

    Science.gov (United States)

    Yilmaz, Vuslat; Oflazer, Piraye; Aysal, Fikret; Parman, Yeşim G; Direskeneli, Haner; Deymeer, Feza; Saruhan-Direskeneli, Güher

    2015-06-01

    B cells from myasthenia gravis (MG) patients with autoantibodies (Aab) against acetylcholine receptor (AChR), muscle-specific kinase (MuSK) or with no detectable Aab were investigated as cytokine producing cells in this study. B cells were evaluated for memory phenotypes and expressions of IL-10, IL-6 and IL-12A. Induced productions of IL-10, IL-6, IL-12p40, TNF-α and LT from isolated B cells in vitro were measured by immunoassays. MG patients receiving immunosuppressive treatment had higher proportions of memory B cells compared with healthy controls and untreated patients. With CD40 stimulation MG patients produced significantly lower levels of IL-10, IL-6. With CD40 and B cell receptor stimulation of B cells, TNF-α production also decreased in addition to these cytokines. The lower levels of these cytokine productions were not related to treatment. Our results confirm a disturbance of B cell subpopulations in MG subgroups on immunosuppressive treatment. B cell derived IL-10, IL-6 and TNF-α are down-regulated in MG, irrespective of different antibody productions. Ineffective cytokine production by B cells may be a susceptibility factor in dysregulation of autoimmune Aab production.

  18. Conventional NK cells can produce IL-22 and promote host defense in Klebsiella pneumoniae pneumonia.

    Science.gov (United States)

    Xu, Xin; Weiss, Ido D; Zhang, Hongwei H; Singh, Satya P; Wynn, Thomas A; Wilson, Mark S; Farber, Joshua M

    2014-02-15

    It was reported that host defense against pulmonary Klebsiella pneumoniae infection requires IL-22, which was proposed to be of T cell origin. Supporting a role for IL-22, we found that Il22(-/-) mice had decreased survival compared with wild-type mice after intratracheal infection with K. pneumoniae. Surprisingly, however, Rag2(-/-) mice did not differ from wild-type mice in survival or levels of IL-22 in the lungs postinfection with K. pneumoniae. In contrast, K. pneumoniae-infected Rag2(-/-)Il2rg(-/-) mice failed to produce IL-22. These data suggested a possible role for NK cells or other innate lymphoid cells in host defense and production of IL-22. Unlike NK cell-like innate lymphoid cells that produce IL-22 and display a surface phenotype of NK1.1(-)NKp46(+)CCR6(+), lung NK cells showed the conventional phenotype, NK1.1(+)NKp46(+)CCR6(-). Mice depleted of NK cells using anti-asialo GM1 showed decreased survival and higher lung bacterial counts, as well as increased dissemination of K. pneumoniae to blood and liver, compared with control-treated mice. NK cell depletion also led to decreased production of IL-22 in the lung. Within 1 d postinfection, although there was no increase in the number of lung NK cells, a subset of lung NK cells became competent to produce IL-22, and such cells were found in both wild-type and Rag2(-/-) mice. Our data suggest that, during pulmonary infection of mice with K. pneumoniae, conventional NK cells are required for optimal host defense, which includes the production of IL-22.

  19. In vitro cell cultures obtained from different explants of Corylus avellana produce Taxol and taxanes

    Directory of Open Access Journals (Sweden)

    Cavalli Francesca

    2006-12-01

    Full Text Available Abstract Background Taxol is an effective antineoplastic agent, originally extracted from the bark of Taxus brevifolia with a low yield. Many attempts have been made to produce Taxol by chemical synthesis, semi-synthesis and plant tissue cultures. However, to date, the availability of this compound is not sufficient to satisfy the commercial requirements. The aim of the present work was to produce suspension cell cultures from plants not belonging to Taxus genus and to verify whether they produced Taxol and taxanes. For this purpose different explants of hazel (Corylus avellana species were used to optimize the protocol for inducing in vitro callus, an undifferentiated tissue from which suspension cell cultures were established. Results Calli were successfully induced from stems, leaves and seeds grown in various hormone concentrations and combinations. The most suitable callus to establish suspension cell cultures was obtained from seeds. Media recovered from suspension cell cultures contained taxanes, and showed antiproliferative activity on human tumour cells. Taxol, 10-deacetyltaxol and 10-deacetylbaccatin III were the main taxanes identified. The level of Taxol recovered from the media of hazel cultures was similar to that found in yew cultures. Moreover, the production of taxanes in hazel cell cultures increased when elicitors were used. Conclusion Here we show that hazel cell cultures produce Taxol and taxanes under controlled conditions. This result suggests that hazel possesses the enzymes for Taxol production, which until now was considered to be a pathway particular to Taxus genus. The main benefit of producing taxanes through hazel cell cultures is that hazel is widely available, grows at a much faster rate in vivo, and is easier to cultivate in vitro than yew. In addition, the production of callus directly from hazel seeds shortens the culture time and minimizes the probability of contamination. Therefore, hazel could become a

  20. A cell sorting protocol for selecting high-producing sub-populations of Sf9 and High Five™ cells.

    Science.gov (United States)

    Vidigal, João; Dias, Mafalda M; Fernandes, Fabiana; Patrone, Marco; Bispo, Cláudia; Andrade, Cláudia; Gardner, Rui; Carrondo, Manuel J T; Alves, Paula M; Teixeira, Ana P

    2013-12-01

    Insect cell lines such as Sf9 and High Five™ have been widely used to produce recombinant proteins mostly by the lytic baculovirus vector system. We have recently established an expression platform in Sf9 cells using a fluorescence-based recombinase mediated cassette exchange (RMCE) strategy which has similar development timelines but avoids baculovirus infection. To expedite cell engineering efforts, a robust fluorescence-activated cell sorting (FACS) protocol optimized for insect cells was developed here. The standard sorting conditions used for mammalian cells proved to be unsuitable, resulting in post-sorting viabilities below 10% for both cell lines. We found that the extreme sensitivity to the shear stress displayed by Sf9 and High Five™ cells was the limiting factor, and using Pluronic F-68 in the cell suspension could increase post-sorting viabilities in a dose dependent manner. The newly developed protocol was then used to sort stable populations of both cell lines tagged with a DsRed-expressing cassette. Before sorting, the average fluorescence intensity of the Sf9 cell population was 3-fold higher than that of the High Five™ cell population. By enriching with the 10% strongest DsRed-fluorescent cells, the productivity of both cell populations could be successfully improved. The established sorting protocol potentiates the use of RMCE technology for recombinant protein production in insect cells.

  1. Chitosan Conduit for Peripheral Nerve Regeneration

    Institute of Scientific and Technical Information of China (English)

    1999-01-01

    Chitosan, the N-deacetylated form of chitin, has good biocompatibility and biodegradability.This paper investigates the feasibility of using chitosan conduits for peripheral nerve regeneration.Cell culture experiments were used to test the material's cytotoxicity and affinity to nerve cells.Conduit implantation experiments were used to study the degradation of the material and the regeneration of injured sciatic nerves.The primary results indicate that chitosan has good mechanical properties, biocompatibility, and biodegradability and it may be a promising biomaterial for peripheral nerve regeneration.

  2. Host cell capable of producing enzymes useful for degradation of lignocellulosic material

    Science.gov (United States)

    Los, Alrik Pieter; Sagt, Cornelis Maria Jacobus; Schooneveld-Bergmans, Margot Elisabeth Francoise; Damveld, Robbertus Antonius

    2015-08-18

    The invention relates to a host cell comprising at least four different heterologous polynucleotides chosen from the group of polynucleotides encoding cellulases, hemicellulases and pectinases, wherein the host cell is capable of producing the at least four different enzymes chosen from the group of cellulases, hemicellulases and pectinases, wherein the host cell is a filamentous fungus and is capable of secretion of the at least four different enzymes. This host cell can suitably be used for the production of an enzyme composition that can be used in a process for the saccharification of cellulosic material.

  3. Polymeric Nerve Conduits with Contact Guidance Cues Used in Nerve Repair

    Institute of Scientific and Technical Information of China (English)

    G DAI; X NIU; J YIN

    2016-01-01

    In the modern life, the nerve injury frequently happens due to mechanical, chemical or thermal accidents. In the trivial injuries, the peripheral nerves can regenerate on their own; however, in most of the cases the clinical treatments are required, where relatively large nerve injury gaps are formed. Currently, the nerve repair can be accomplished by direct suture when the injury gap is not too large;while the autologous nerve graft working as the gold standard of peripheral nerve injury treatment for nerve injuries with larger gaps. However, the direct suture is limited by heavy tension at the suture sites, and the autologous nerve graft also has the drawbacks of donor site morbidity and insufifcient donor tissue. Recently, artiifcial nerve conduits have been developed as an alternative for clinical nerve repair to overcome the limitations associated with the above treatments. In order to further improve the efifciency of nerve conduits, various guidance cues are incorporated, including physical cues, biochemical signals, as well as support cells. First, this paper reviewed the contact guidance cues applied in nerve conduits, such as lumen ifllers, multi-channels and micro-patterns on the inner surface. Then, the paper focused on the polymeric nerve conduits with micro inner grooves. The polymeric nerve conduits were fabricated using the phase inversion-based ifber spinning techniques. The smart spinneret with grooved die was designed in the spinning platform, while different spinning conditions, including flow rates, air-gap distances, and polymer concentrations, were adjusted to investigate the inlfuence of fabrication conditions on the geometry of nerve conduits. The inner groove size in the nerve conduits can be precisely controlled in our hollow ifber spinning process, which can work as the efifcient contact guidance cue for nerve regeneration.

  4. Hox6 genes modulate in vitro differentiation of mESCs to insulin-producing cells.

    Science.gov (United States)

    Larsen, Brian M; Marty-Santos, Leilani; Newman, Micaleah; Lukacs, Derek T; Spence, Jason R; Wellik, Deneen M

    2016-10-01

    The differentiation of glucose-responsive, insulin-producing cells from ESCs in vitro is promising as a cellular therapy for the treatment of diabetes, a devastating and common disease. Pancreatic β-cells are derived from the endoderm in vivo and therefore most current protocols attempt to generate a pure population of first endoderm, then pancreas epithelium, and finally insulin-producing cells. Despite this, differentiation protocols result in mixed populations of cells that are often poorly defined, but also contain mesoderm. Using an in vitro mESC-to-β cell differentiation protocol, we show that expression of region-specific Hox genes is induced. We also show that the loss of function of the Hox6 paralogous group, genes expressed only in the mesenchyme of the pancreas (not epithelium), affect the differentiation of insulin-producing cells in vitro. This work is consistent with the important role for these mesoderm-specific factors in vivo and highlights contribution of supporting mesenchymal cells in in vitro differentiation.

  5. GRP nerves in pig antrum

    DEFF Research Database (Denmark)

    Holst, J J; Poulsen, Steen Seier

    1987-01-01

    We extracted gastrin-releasing peptide (GRP) and its C-terminal decapeptide corresponding to 6.4 and 6.8 pmol/g from pig antrum mucosa. By immunohistochemistry GRP was localized to mucosal, submucosal, and myenteric nerve fibers. A few nerve cell bodies were also identified. Using isolated perfused...... pig antrum with intact vagal innervation, we found concomitant, atropine-resistant release of GRP and gastrin during electrical stimulation of the vagal nerves. Intra-arterial GRP at 10(-11)-10(-10) mol/l caused up to fivefold, dose-dependent increases in gastrin secretion; higher doses were less...... response to GRP and abolished the effect of vagal stimulation. The available evidence strongly suggests that GRP nerves are responsible for the stimulatory vagal effects on gastrin secretion in the pig....

  6. Liver sinusoidal endothelial cells induce immunosuppressive IL-10-producing Th1 cells via the Notch pathway.

    Science.gov (United States)

    Neumann, Katrin; Rudolph, Christine; Neumann, Christian; Janke, Marko; Amsen, Derk; Scheffold, Alexander

    2015-07-01

    Under homeostasis, liver sinusoidal endothelial cells (LSECs) shift intrahepatic T-cell responses towards tolerance. However, the role of LSECs in the regulation of T-cell-induced liver inflammation is less clear. Here, we studied the capacity of LSECs to modulate pro-inflammatory Th1-cell differentiation in mice. Using in vitro co-culture systems and subsequent cytokine analysis, we showed that LSECs induced high amounts of the anti-inflammatory cytokine IL-10 in developing Th1 cells. These LSEC-stimulated Th1 cells had no pro-inflammatory capacity in vivo but instead actively suppressed an inflammatory Th1-cell-induced delayed-type hypersensitivity reaction. Blockage of IL-10 signaling in vivo inhibited immunosuppressive activity of LSEC-stimulated Th1 cells. We identified the Notch pathway as a mechanism how LSECs trigger IL-10 expression in Th1 cells. LSECs expressed high levels of the Delta-like and Jagged family of Notch ligands and induced expression of the Notch target genes hes-1 and deltex-1 in Th1 cells. Blockade of Notch signaling selectively inhibited IL-10 induction in Th1 cells by LSECs. Our findings suggest that LSEC-induced IL-10 expression in Th1 cells via the Notch pathway may contribute to the control of hepatic inflammatory immune responses by induction of a self-regulatory mechanism in pro-inflammatory Th1 cells.

  7. Immobilized gellan sulfate surface for cell adhesion and multiplication: development of cell-hybrid biomaterials using self-produced fibronectin.

    Science.gov (United States)

    Miyamoto, Keiichi; Kanemoto, Akiko; Hashimoto, Kenichi; Tokita, Masayuki; Komai, Takashi

    2002-04-08

    A new concept for cell-hybrid biomaterial is proposed in which human unbilical vein endothelial cells (HUVEC) are adhered to an immobilized gellan sulfate (GS) surface. Extra domain A containing fibronectin (EDA(+)FN) released from HUVEC is necessary for cell adhesion and multiplication. The material design in this study is based on these self-released cell adhesion proteins. The interaction between GS and EDA(+)FN was evaluated using the affinity constant (KA); the value obtained was 1.03x10(8) (M(-1)). These results suggest that the adhesion of HUVEC to GS may be supported by the adhesion of EDA(+)FN to GS. We also found that this new material adheres to HUVEC, allowing the reintroduction of EDA(+)FN, which is self-produced by the cell. This material is relatively easy to produce, not requiring the usual coating of adhesion proteins in pretreatment.

  8. [A case of alpha-cell nesidioblastosis and hyperplasia with multiple glucagon-producing endocrine cell tumor of the pancreas].

    Science.gov (United States)

    Kang, Huapyong; Kim, Sewha; Lim, Tae Seop; Lee, Hye Won; Choi, Heun; Kang, Chang Moo; Kim, Ho Guen; Bang, Seungmin

    2014-04-01

    Nesidioblastosis is a term used to describe pathologic overgrowth of pancreatic islet cells. It also means maldistribution of islet cells within the ductules of exocrine pancreas. Generally, nesidioblastosis occurs in beta-cell and causes neonatal hyperinsulinemic hypoglycemia or adult noninsulinoma pancreatogenous hypoglycemia syndrome. Alpha-cell nesidioblastosis and hyperplasia is an extremely rare disorder. It often accompanies glucagon-producing marco- and mircoadenoma without typical glucagonoma syndrome. A 35-year-old female was referred to our hospital with recurrent acute pancreatitis. On radiologic studies, 1.5 cm sized mass was noted in pancreas tail. Cytological evaluation with EUS-fine-needle aspiration suggested serous cystadenoma. She received distal pancreatectomy. The histologic examination revealed a 1.7 cm sized neuroendocrine tumor positive for immunohistochemical staining with glucagon antibody. Multiple glucagon-producing micro endocrine cell tumors were scattered next to the main tumor. Additionally, diffuse hyperplasia of pancreatic islets and ectopic proliferation of islet cells in centroacinar area, findings compatible to nesidioblastosis, were seen. These hyperplasia and almost all nesidioblastic cells were positive for glucagon immunochemistry. Even though serum glucagon level still remained higher than the reference value, she has been followed-up without any evidence of recurrence or hormone related symptoms. Herein, we report a case of alpha-cell nesidioblastosis and hyperplasia combined with glucagon-producing neuroendocrine tumor with literature review.

  9. Correlation between standard plate count and somatic cell count milk quality results for Wisconsin dairy producers.

    Science.gov (United States)

    Borneman, Darand L; Ingham, Steve

    2014-05-01

    The objective of this study was to determine if a correlation exists between standard plate count (SPC) and somatic cell count (SCC) monthly reported results for Wisconsin dairy producers. Such a correlation may indicate that Wisconsin producers effectively controlling sanitation and milk temperature (reflected in low SPC) also have implemented good herd health management practices (reflected in low SCC). The SPC and SCC results for all grade A and B dairy producers who submitted results to the Wisconsin Department of Agriculture, Trade, and Consumer Protection, in each month of 2012 were analyzed. Grade A producer SPC results were less dispersed than grade B producer SPC results. Regression analysis showed a highly significant correlation between SPC and SCC, but the R(2) value was very small (0.02-0.03), suggesting that many other factors, besides SCC, influence SPC. Average SCC (across 12 mo) for grade A and B producers decreased with an increase in the number of monthly SPC results (out of 12) that were ≤ 25,000 cfu/mL. A chi-squared test of independence showed that the proportion of monthly SCC results >250,000 cells/mL varied significantly depending on whether the corresponding SPC result was ≤ 25,000 or >25,000 cfu/mL. This significant difference occurred in all months of 2012 for grade A and B producers. The results suggest that a generally consistent level of skill exists across dairy production practices affecting SPC and SCC.

  10. Immunohistochemical, ultrastructural and functional analysis of axonal regeneration through peripheral nerve grafts containing Schwann cells expressing BDNF, CNTF or NT3.

    Directory of Open Access Journals (Sweden)

    Maria João Godinho

    Full Text Available We used morphological, immunohistochemical and functional assessments to determine the impact of genetically-modified peripheral nerve (PN grafts on axonal regeneration after injury. Grafts were assembled from acellular nerve sheaths repopulated ex vivo with Schwann cells (SCs modified to express brain-derived neurotrophic factor (BDNF, a secretable form of ciliary neurotrophic factor (CNTF, or neurotrophin-3 (NT3. Grafts were used to repair unilateral 1 cm defects in rat peroneal nerves and 10 weeks later outcomes were compared to normal nerves and various controls: autografts, acellular grafts and grafts with unmodified SCs. The number of regenerated βIII-Tubulin positive axons was similar in all grafts with the exception of CNTF, which contained the fewest immunostained axons. There were significantly lower fiber counts in acellular, untransduced SC and NT3 groups using a PanNF antibody, suggesting a paucity of large caliber axons. In addition, NT3 grafts contained the greatest number of sensory fibres, identified with either IB4 or CGRP markers. Examination of semi- and ultra-thin sections revealed heterogeneous graft morphologies, particularly in BDNF and NT3 grafts in which the fascicular organization was pronounced. Unmyelinated axons were loosely organized in numerous Remak bundles in NT3 grafts, while the BDNF graft group displayed the lowest ratio of umyelinated to myelinated axons. Gait analysis revealed that stance width was increased in rats with CNTF and NT3 grafts, and step length involving the injured left hindlimb was significantly greater in NT3 grafted rats, suggesting enhanced sensory sensitivity in these animals. In summary, the selective expression of BDNF, CNTF or NT3 by genetically modified SCs had differential effects on PN graft morphology, the number and type of regenerating axons, myelination, and locomotor function.

  11. Spatially- and temporally-controlled postnatal p53 knockdown cooperates with embryonic Schwann cell precursor Nf1 gene loss to promote malignant peripheral nerve sheath tumor formation.

    Science.gov (United States)

    Hirbe, Angela C; Dahiya, Sonika; Friedmann-Morvinski, Dinorah; Verma, Inder M; Clapp, D Wade; Gutmann, David H

    2016-02-16

    Malignant peripheral nerve sheath tumors (MPNSTs) are highly aggressive sarcomas that arise sporadically or in association with the Neurofibromatosis type 1 (NF1) cancer predisposition syndrome. In individuals with NF1, MPNSTs are hypothesized to arise from Nf1-deficient Schwann cell precursor cells following the somatic acquisition of secondary cooperating genetic mutations (e.g., p53 loss). To model this sequential genetic cooperativity, we coupled somatic lentivirus-mediated p53 knockdown in the adult right sciatic nerve with embryonic Schwann cell precursor Nf1 gene inactivation in two different Nf1 conditional knockout mouse strains. Using this approach, ~60% of mice with Periostin-Cre-mediated Nf1 gene inactivation (Periostin-Cre; Nf1(flox/flox) mice) developed tumors classified as low-grade MPNSTs following p53 knockdown (mean, 6 months). Similarly, ~70% of Nf1+/- mice with GFAP-Cre-mediated Nf1 gene inactivation (GFAP-Cre; Nf1(flox/null) mice) developed low-grade MPNSTs following p53 knockdown (mean, 3 months). In addition, wild-type and Nf1+/- mice with GFAP-Cre-mediated Nf1 loss develop MPNSTs following somatic p53 knockout with different latencies, suggesting potential influences of Nf1+/- stromal cells in MPNST pathogenesis. Collectively, this new MPNST model system permits the analysis of somatically-acquired events as well as tumor microenvironment signals that potentially cooperate with Nf1 loss in the development and progression of this deadly malignancy.

  12. Platelet Lysates Produced from Expired Platelet Concentrates Support Growth and Osteogenic Differentiation of Mesenchymal Stem Cells

    OpenAIRE

    Sandra Mjoll Jonsdottir-Buch; Ramona Lieder; Olafur Eysteinn Sigurjonsson

    2013-01-01

    BACKGROUND: Mesenchymal stem cells are promising candidates in regenerative cell therapy. Conventional culture methods involve the use of animal substances, specifically fetal bovine serum as growth supplement. Since the use of animal-derived products is undesirable for human applications, platelet lysates produced from human platelets are an attractive alternative. This is especially true if platelet lysates from already approved transfusion units at blood banks can be utilized. The purpose ...

  13. Chinese Experts Successfully Produced Transgenic Animals from Haploid Embryonic Stem Cells

    Institute of Scientific and Technical Information of China (English)

    2012-01-01

    Individual animals produced by haploid stem cells are ideal models for studying recessive genes. Hap- loid stem cells not only can maintain haploidy, but also are capable of replicating themselves infinitely. Modified genes can be passed on to future generations through genetic engineering of haploid embryonic stem cells, which thus avoids the germlinechimerism caused by other transgenic methods and greatly im- proves the analysis efficiency of the function of gene modification. However, natural haploids are only re- stricted to germline cells in mammals. Currently in mammals, only the embryonic stem cells in rats and mice can be used as the carrier of gene modification, but the embryonic stem cells of other mammals, in- eluding primates, cannot guarantee germline transmission, which has seriously hindered the establishment of disease models by using these species.

  14. A Rapid Culture Technique Produces Functional Dendritic-Like Cells from Human Acute Myeloid Leukemia Cell Lines

    Directory of Open Access Journals (Sweden)

    Jian Ning

    2011-01-01

    Full Text Available Most anti-cancer immunotherapeutic strategies involving dendritic cells (DC as vaccines rely upon the adoptive transfer of DC loaded with exogenous tumour-peptides. This study utilized human acute myeloid leukemia (AML cells as progenitors from which functional dendritic-like antigen presenting cells (DLC were generated, that constitutively express tumour antigens for recognition by CD8+ T cells. DLC were generated from AML cell lines KG-1 and MUTZ-3 using rapid culture techniques and appropriate cytokines. DLC were evaluated for their cell-surface phenotype, antigen uptake and ability to stimulate allogeneic responder cell proliferation, and production of IFN-γ; compared with DC derived from normal human PBMC donors. KG-1 and MUTZ-3 DLC increased expression of CD80, CD83, CD86, and HLA-DR, and MUTZ-3 DLC downregulated CD14 and expressed CD1a. Importantly, both KG-1 and MUTZ-3-derived DLC promoted proliferation of allogeneic responder cells more efficiently than unmodified cells; neither cells incorporated FITC-labeled dextran, but both stimulated IFN-γ production from responding allogeneic CD8+ T cells. Control DC produced from PBMC using the FastDC culture also expressed high levels of critical cell surface ligands and demonstrated good APC function. This paper indicates that functional DLC can be cultured from the AML cell lines KG-1 and MUTZ-3, and FastDC culture generates functional KG-1 DLC.

  15. High-throughput sorting of the highest producing cell via a transiently protein-anchored system.

    Directory of Open Access Journals (Sweden)

    Kuo-Hsiang Chuang

    Full Text Available Developing a high-throughput method for the effecient selection of the highest producing cell is very important for the production of recombinant protein drugs. Here, we developed a novel transiently protein-anchored system coupled with fluorescence activated cell sorting (FACS for the efficient selection of the highest producing cell. A furin cleavage peptide (RAKR was used to join a human anti-epithelial growth factor antibody (αEGFR Ab and the extracellular-transmembrane-cytosolic domains of the mouse B7-1 antigen (B7. The furin inhibitor can transiently switch secreted αEGFR Ab into a membrane-anchored form. After cell sorting, the level of membrane αEGFR Ab-RAKR-B7 is proportional to the amount of secreted αEGFR Ab in the medium. We further selected 23 αEGFR Ab expressing cells and demonstrated a high correlation (R2 = 0.9165 between the secretion level and surface expression levels of αEGFR Ab. These results suggested that the novel transiently protein-anchored system can easily and efficiently select the highest producing cells, reducing the cost for the production of biopharmaceuticals.

  16. Effect of Acteoside as a Cell Protector to Produce a Cloned Dog

    Science.gov (United States)

    Kim, Keun Jung; Kim, Eun Young; Kim, Dong-hee; Lee, Bo Myeong; Han, Kil Woo; Park, Kang-Sun; Lee, Kyung-Bon; Kim, Min Kyu

    2016-01-01

    Somatic cell nuclear transfer (SCNT) is a well-known laboratory technique. The principle of the SCNT involves the reprogramming a somatic nucleus by injecting a somatic cell into a recipient oocyte whose nucleus has been removed. Therefore, the nucleus donor cells are considered as a crucial factor in SCNT. Cell cycle synchronization of nucleus donor cells at G0/G1 stage can be induced by contact inhibition or serum starvation. In this study, acteoside, a phenylpropanoid glycoside compound, was investigated to determine whether it is applicable for inducing cell cycle synchronization, cytoprotection, and improving SCNT efficiency in canine fetal fibroblasts. Primary canine fetal fibroblasts were treated with acteoside (10, 30, 50 μM) for various time periods (24, 48 and 72 hours). Cell cycle synchronization at G0/G1 stage did not differ significantly with the method of induction: acteoside treatment, contact inhibition or serum starvation. However, of these three treatments, serum starvation resulted in significantly increased level of reactive oxygen species (ROS) (99.5 ± 0.3%) and apoptosis. The results also revealed that acteoside reduced ROS and apoptosis processes including necrosis in canine fetal fibroblasts, and improved the cell survival. Canine fetal fibroblasts treated with acteoside were successfully arrested at the G0/G1 stage. Moreover, the reconstructed embryos using nucleus donor cells treated with acteoside produced a healthy cloned dog, but not the embryos produced using nucleus donor cells subjected to contact inhibition. In conclusion, acteoside induced cell cycle synchronization of nucleus donor cells would be an alternative method to improve the efficiency of canine SCNT because of its cytoprotective effects. PMID:27428333

  17. De Novo Formation of Insulin-Producing “Neo-β Cell Islets” from Intestinal Crypts

    Directory of Open Access Journals (Sweden)

    Yi-Ju Chen

    2014-03-01

    Full Text Available The ability to interconvert terminally differentiated cells could serve as a powerful tool for cell-based treatment of degenerative diseases, including diabetes mellitus. To determine which, if any, adult tissues are competent to activate an islet β cell program, we performed an in vivo screen by expressing three β cell “reprogramming factors” in a wide spectrum of tissues. We report that transient intestinal expression of these factors—Pdx1, MafA, and Ngn3 (PMN—promotes rapid conversion of intestinal crypt cells into endocrine cells, which coalesce into “neoislets” below the crypt base. Neoislet cells express insulin and show ultrastructural features of β cells. Importantly, intestinal neoislets are glucose-responsive and able to ameliorate hyperglycemia in diabetic mice. Moreover, PMN expression in human intestinal “organoids” stimulates the conversion of intestinal epithelial cells into β-like cells. Our results thus demonstrate that the intestine is an accessible and abundant source of functional insulin-producing cells.

  18. Bone Marrow Homing Enriches Stem Cells Responsible for Neogenesis of Insulin-Producing Cells, While Radiation Decreases Homing Efficiency.

    Science.gov (United States)

    Goldenberg-Cohen, Nitza; Iskovich, Svetlana; Askenasy, Nadir

    2015-10-01

    Small-sized adult bone marrow cells isolated by counterflow centrifugal elutriation and depleted of lineage markers (Fr25lin(-)) have the capacity to differentiate into insulin-producing cells and stabilize glycemic control. This study assessed competitive migration of syngeneic stem cells to the bone marrow and islets in a murine model of chemical diabetes. VLA-4 is expressed in ∼ 25% of these cells, whereas CXCR4 is not detected, however, it is transcriptionally upregulated (6-fold). The possibility to enrich stem cells by a bone marrow homing (BM-H) functional assay was assessed in sequential transplants. Fr25lin(-) cells labeled with PKH26 were grafted into primary myeloablated recipients, and mitotically quiescent Fr25lin(-)PKH(bright) cells were sorted from the bone marrow after 2 days. The contribution of bone marrow-homed stem cells was remarkably higher in secondary recipients compared to freshly elutriated cells. The therapeutic efficacy was further increased by omission of irradiation in the secondary recipients, showing a 25-fold enrichment of islet-reconstituting cells by the bone marrow homing assay. Donor cells identified by the green fluorescent protein (GFP) and a genomic marker in sex-mismatched transplants upregulated PDX-1 and produced proinsulin, affirming the capacity of BM-H cells to convert in the injured islets. There was no evidence of transcriptional priming of freshly elutriated subsets to express PDX-1, insulin, and other markers of endocrine progenitors, indicating that the bone marrow harbors stem cells with versatile differentiation capacity. Affinity to the bone marrow can be used to enrich stem cells for pancreatic regeneration, and reciprocally, conditioning reduces the competitive incorporation in the injured islets.

  19. Effects of nerve growth factor and basic fibroblast growth factor dual gene modification on rat bone marrow mesenchymal stem cell differentiation into neuron-like cells in vitro.

    Science.gov (United States)

    Hu, Yang; Zhang, Yan; Tian, Kang; Xun, Chong; Wang, Shouyu; Lv, Decheng

    2016-01-01

    Recent studies regarding regenerative medicine have focused on bone marrow mesenchymal stem cells (BMSCs), which have the potential to undergo neural differentiation, and may be transfected with specific genes. BMSCs can differentiate into neuron‑like cells in certain neurotropic circumstances in vitro. Basic fibroblast growth factor (bFGF) and nerve growth factor (NGF) are often used to induce neural differentiation in BMSCs in vitro. However, previous studies regarding their combined actions are insufficient. The present study is the first, to the best of our knowledge, to thoroughly assess the enhancement of neural differentiation of BMSCs following transfection with bFGF and NGF. Sprague‑Dawley (SD) rat BMSCs were separated through whole bone marrow adherence, and were then passaged to the third generation. The cells were subsequently divided into five groups: The control group, which consisted of untransfected BMSCs; the plv‑blank‑transfected BMSCs group; the plv‑bFGF‑transfected BMSCs group; the plv‑NGF‑transfected BMSCs group; and the plv‑NGF‑bFGF co‑transfected BMSCs group. Cell neural differentiation was characterized in terms of stem cell molecular expression, and the neuronal morphology and expression of neural‑like molecules was detected in each of the groups. A total of 72 h post‑transfection, the expression levels of neuron‑specific enolase, glial fibrillary acidic protein, and nestin protein, were higher in the co‑transfected group, as compared with the other groups, the expression levels of β‑tubulin III were also increased in the co‑transfected cells, thus suggesting the maturation of differentiated neuron‑like cells. Furthermore, higher neuronal proliferation was observed in the co‑transfected group, as compared with the other groups at passages 2, 4, 6 and 8. Western blotting demonstrated that the transfected groups exhibited a simultaneous increase in phosphorylation of the AKT and extracellular signal

  20. Growth-promoting activity of Hominis Placenta extract on regenerating sciatic nerve

    Institute of Scientific and Technical Information of China (English)

    Tae-beom SEO; Dong-hee KIM; Seung-kiel PARK; Deok-chun YANG; Uk NAMGUNG; In-sun HAN; Jin-hwan YOON; In-chan SEOL; Yun-sik KIM; Hyun-kyung JO; Joung-jo AN; Kwon-eui HONG; Young-bae SEO

    2006-01-01

    Aim: Extract of Hominis Placenta (HP) has been used in oriental medicine as an agent for improving physiological function. The present study was conducted to investigate whether HP treatment in an experimental sciatic nerve injury animal model produces growth-promoting effects on regenerating peripheral nerve fibers after injury. Methods: After HP was injected into a sciatic nerve injury site, changes in protein levels were analyzed in the regenerating nerve area by Western blotting and immunofluorescence staining analyses. For quantitative assessment of axonal regeneration, a retrograde tracing technique was used to identify the neuronal cell bodies corresponding to regenerating axons, and the extent of neurite outgrowth in cultured dorsal root ganglia (DRG) sensory neurons prepared from animals that had experienced a sciatic nerve crush injury 7 d before neuron collection was analyzed. Results: Induction levels of axonal growth-associated protein (GAP-43) in the injured sciatic nerves were elevated by HP treatment. HP treatment also upregulated cell division cycle 2 (Cdc2) protein levels in the distal stump of the injured sciatic nerve. Induced Cdc2 protein was detected in Schwann cells, suggesting that Cdc2 kinase activity may be involved in the growth-promoting activity of regenerating axons via Schwann cell proliferation. Cell body measurement by retrograde tracing indicated that HP treatment produced significant increases in regenerating motor axons. Finally, HP treatment of cultured DRG sensory neurons significantly increased neurite arborization and elongation.Conclusion: HP promotes the regeneration of injured sciatic axons by upregulating the synthesis of regeneration-related protein factors such as GAP-43 and Cdc2.

  1. Platelet lysates produced from expired platelet concentrates support growth and osteogenic differentiation of mesenchymal stem cells.

    Directory of Open Access Journals (Sweden)

    Sandra Mjoll Jonsdottir-Buch

    Full Text Available BACKGROUND: Mesenchymal stem cells are promising candidates in regenerative cell therapy. Conventional culture methods involve the use of animal substances, specifically fetal bovine serum as growth supplement. Since the use of animal-derived products is undesirable for human applications, platelet lysates produced from human platelets are an attractive alternative. This is especially true if platelet lysates from already approved transfusion units at blood banks can be utilized. The purpose of this study was to produce human platelet lysates from expired, blood bank-approved platelet concentrates and evaluate their use as growth supplement in the culture of mesenchymal stem cells. METHODOLOGY/PRINCIPAL FINDINGS: In this study, bone marrow-derived mesenchymal stem cells were cultured with one of three culture supplements; fetal bovine serum, lysates from freshly prepared human platelet concentrates, or lysates from expired human platelet concentrates. The effects of these platelet-derived culture supplements on basic mesenchymal stem cell characte