Résumé
Nogo-B receptor (NgBR) is a specific receptor of Nogo-B, a member of reticulon 4 protein family. It is widely expressed in many tissues and mainly located in cell membrane and endoplasmic reticulum. Previous studies have revealed that NgBR is involved in a variety of physiological and pathophysiological processes, such as dolichol synthesis, lipid metabolism, cholesterol trafficking, insulin resistance, vascular remodeling and angiogenesis, tumorigenesis and nervous system diseases. Further studies on the molecular characteristics and biological function of NgBR might be of great significance to understand its role in diverse diseases and provide possible clinical strategies for the treatment of diseases.
Sujets)
Protéines de transport/métabolisme , Réticulum endoplasmique/métabolisme , Métabolisme lipidique , Protéines Nogo/métabolisme , Récepteurs de surface cellulaire/métabolismeRésumé
Regeneration of injured peripheral nerves is an extremely complex process. Nogo-A (neurite outgrowth inhibitor-A) inhibits axonal regeneration by interacting with Nogo receptor in the myelin sheath of the central nervous system (CNS). The aim of this study was to investigate the effects of Nogo-A and its receptor on the repair of sciatic nerve injury in rats. Sprague-Dawley rats (n=96) were randomly divided into 4 groups: control group (control), sciatic nerve transection group (model), immediate repair group (immediate repair), and delayed repair group (delayed repair). The rats were euthanized 1 week and 6 weeks after operation. The injured end tissues of the spinal cord and sciatic nerve were obtained. The protein expressions of Nogo-A and Nogo-66 receptor (NgR) were detected by immunohistochemistry. The protein expressions of Nogo-A, NgR, and Ras homolog family member A (RhoA) were detected by western blot. At 1 week after operation, the pathological changes in the immediate repaired group were less, and the protein expressions of Nogo-A, NgR, and RhoA in the spinal cord and sciatic nerve tissues were decreased (P<0.05) compared with the model group. After 6 weeks, the pathological changes in the immediate repair group and the delayed repair group were alleviated and the protein expressions decreased (P<0.05). The situation of the immediate repair group was better than that of the delayed repair group. Our data suggest that the expression of Nogo-A and its receptor increased after sciatic nerve injury, indicating that Nogo-A and its receptor play an inhibitory role in the repair process of sciatic nerve injury in rats.
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Animaux , Rats , Récepteurs de surface cellulaire , Protéines de la myéline , Nerf ischiatique , Rat Sprague-Dawley , Protéines liées au GPI , Protéines Nogo , Régénération nerveuseRésumé
<p><b>OBJECTIVE</b>To study the effects of Jisuikang (Chinese characters) on Nogo-NgR gene expression, and to explore the protective effects and mechanism of Jisuikang (Chinese characters) on spinal cord injury in rats.</p><p><b>METHODS</b>One hundred eighty female rats were randomly assigned to 6 groups(30 rats per group). Sham group: T10 lamina was resected only and spinal cord was untreated. Model group: spine cord injury (SCI) was created with a modified impinger of Allen's by impacting on the T10 spinal cord. Prednisolone group: Prednisolone (0.06 g/kg) was given by intragastric administration at a time interval of 24 hours after operation. The Jisuikang (Chinese characters) high, moderate and low dose groups: Jisuikang (Chinese characters) was supplied with different dose (50 g/kg, 25 g/kg, 12.5 g/kg) by intragastric administration in rats after operation,for the first time at 30 min after surgery. Animals were killed 3, 7, 14 days after surgery. The expression levels of Nogo-A and NgR were observed by Western Blot and Real-time PCR.</p><p><b>RESULTS</b>The expression of Nogo-A and NgR was at the basic level at all time points in sham group. Compared with model group, the protein expression levels of Nogo-A and NgR in sham, prednisolone, Jisuikang (Chinese characters) moderate dose groups were statistically significant at all time points (P < 0.05). No difference was found in Jisuikang (Chinese characters) high and low dose groups (P > 0.05). Three days after surgery, the mRNA levels of Nogo-A and NgR in treatment group were significantly lower than that in model group (P < 0.01); 7 days after surgery,Nogo-A and NgR mRNA expression were dramatically upregulated and peaked; 14 days after operation, the expression was decreased, but still significantly higher than that in other treatment groups (P < 0.01). Prednisolone and Jisuikang (Chinese characters) moderate dose groups showed the most significant effects among all groups,but there was no statistically significant difference between two groups (P > 0.05).</p><p><b>CONCLUSION</b>The decoction Jisuikang (Chinese characters) can promote the nerve cell regeneration by regulating Nogo-A and NgR gene expression, activating Nogo- NgR signaling pathways after acute spinal cord injury.</p>
Sujets)
Animaux , Femelle , Rats , Protéines liées au GPI , Génétique , Physiologie , Médecine traditionnelle chinoise , Protéines de la myéline , Génétique , Physiologie , Régénération nerveuse , Protéines Nogo , Récepteur Nogo-1 , Rat Sprague-Dawley , Récepteurs de surface cellulaire , Génétique , Physiologie , Transduction du signal , Traumatismes de la moelle épinière , Traitement médicamenteux , MétabolismeRésumé
Nogo-B is a major family member of the reticulon protein family 4. It is widely expressed in the central nervous system and peripheral tissues, and is mainly located in endoplasmic reticulum and cell membrane. Previous studies have revealed that Nogo-B plays a key role in vascular injury, tissue repair and inflammation process. It also may be critical for apoptosis of tumor cells and central diseases. Further investigation of the molecular characteristics and biological function of Nogo-B might be of great help to understand its role in diverse diseases.
Sujets)
Animaux , Humains , Apoptose , Membrane cellulaire , Physiologie , Réticulum endoplasmique , Physiologie , Inflammation , Protéines de la myéline , Physiologie , Protéines NogoRésumé
<p><b>BACKGROUND</b>One of the reasons for poor neuroregeneration after central nervous system injury is the presence of inhibitory factors such as Nogo. Here, we tested the inhibition of Nogo by RNA interference both in vitro and in vivo, using recombinant adenovirus-mediated transfection of short hairpin RNAs, to explore a new method of treatment for spinal cord injury.</p><p><b>METHODS</b>We designed and cloned two Nogo-specific short hairpin RNAs and an unrelated short hairpin RNA, packaged the clones into adenovirus, and amplified the recombinant virus in 293 cells. We then tested the inhibition of Nogo expression both in vitro in adenovirus-transfected oligodendrocytes and in vivo in spinal cord tissue from adenovirus-transfected spinal cord injury model rats. We tested Nogo expression at the mRNA level by reverse-transcription PCR and at the protein level by Western blotting and immunohistochemistry.</p><p><b>RESULTS</b>In vitro, the two specific Nogo short hairpin RNAs decreased Nogo mRNA expression by 51% and 49%, respectively, compared with Nogo expression in cells transfected with the unrelated control small hairpin RNA (P < 0.005). Similarly, Nogo protein expression decreased by 50% and 48%, respectively (P < 0.005). In vivo, in spinal cord injury model rats, the two specific Nogo short hairpin RNAs decreased Nogo mRNA expression by 45% and 40%, respectively, compared with Nogo expression in spinal cord injury model rats transfected with the unrelated control short hairpin RNA (P < 0.005). The Nogo protein level was similarly decreased.</p><p><b>CONCLUSIONS</b>We were successful in specifically downregulating Nogo at the mRNA and protein levels by adenovirus-mediated delivery of short hairpin RNAs, both in vitro and in vivo. This confirms the effectiveness of RNA interference for the inhibition of Nogo gene expression and the efficiency of using adenovirus for delivery. Thus gene therapy may be an effective treatment for spinal cord injury.</p>
Sujets)
Animaux , Humains , Rats , Adenoviridae , Génétique , Technique de Western , Immunohistochimie , Protéines de la myéline , Génétique , Métabolisme , Protéines Nogo , Interférence par ARN , Petit ARN interférent , Génétique , Rat Sprague-Dawley , Traumatismes de la moelle épinière , ThérapeutiqueRésumé
<p><b>OBJECTIVE</b>To observe the effects of electroacupuncture (EA) on the expressions of Nogo-A and the ultrastructure in the cerebral cortex at different time points after the cerebral ischemia-reperfusion in rats.</p><p><b>METHODS</b>One hundred and thirty male Sprague Dawley (SD) rats were randomly divided into the EA group (n = 30), the sham-EA group (n = 30), the model group (n = 30), the sham-operation group (n = 30), and the blank group (n = 10). The modified ZeaLonga method was used to prepare the left middle cerebral artery occlusion (MCAO) model in the first three groups. After the operation Baihui (DU20) and Dazhui (DU14) were daily needled in the EA group. One inch beside Baihui (DU20) and Dazhui (DU14) were daily needled in the sham-EA group. Rats in the model group were only treated with MCAO ischemia/reperfusion. Rats in the sham-operation group only received surgical wound. No treatment was given to rats in the blank group. The ultrastructures of ischemic cells and the intervention of the Nogo-A expressions were observed using the immunohistochemical staining and the transmission electron microscope 1, 7, and 28 days after EA.</p><p><b>RESULTS</b>(1) In the EA group, the damage of ultrastructures of neurons, gliocytes, and blood brain barrier in the ischemic region was alleviated when compared with that of the sham-EA group and the model group. (2) On the 1st, 7th and 28th day after the cerebral ischemia-reperfusion, the expressions of Nogo-A in the ischemic cortex in the EA group was lower when compared with those in the sham-EA group and the model group at the corresponding time points, showing significant difference (P < 0.05). But there was no statistical difference between the sham-EA group and the model group at the same time point (P > 0.05).</p><p><b>CONCLUSION</b>The mechanism of EA for protecting cerebral ischemia/reperfusion might be closely associated with alleviating the damage on the ultrastructures of brain cells, and down-regulating the expressions of Nogo-A.</p>
Sujets)
Animaux , Mâle , Rats , Points d'acupuncture , Encéphalopathie ischémique , Métabolisme , Anatomopathologie , Thérapeutique , Cortex cérébral , Métabolisme , Électroacupuncture , Protéines de la myéline , Métabolisme , Protéines Nogo , Rat Sprague-Dawley , Lésion d'ischémie-reperfusion , Métabolisme , Anatomopathologie , ThérapeutiqueRésumé
<p><b>OBJECTIVE</b>To study the molecular mechanism of Zuogui Pill (ZGP) and Yougui Pill (YGP) on axonal regeneration in rats with experimental autoimmune encephalomyelitis (EAE).</p><p><b>METHODS</b>EAE rat model was established by bilateral rear pedes subcutaneous injection of antigen made by mixing myelin basic protein (MBP) and complete Freud's adjuvant (CFA) in the volume ratio of 1:1. The pathological changes of axonal injury and regeneration in the brain and the spinal cord were observed on the 14th (the acute stage) and the 28th day (the remission stage) after modeling, with hematoxylin-eosin (HE) staining, silver stain, and immunohistochemical staining. The rats treated with prednisone acetate were taken as controls.</p><p><b>RESULTS</b>Observation under the light microscope with HE staining showed a sleeve-like change in rats' cerebrospinal parenchyma with inflammatory cell infiltration around the small vessels and neuronic denaturation, while silver staining showed excessive tumefaction and abscission of axon, and immunohistochemical analysis showed decreasing of nerve growth factor (NGF) expression at the acute stage of EAE, which was even more remarkable at the remission stage, showing significant difference as compared with the normal control (P<0.05). And the expressions of Nogo A, an axon growth inhibitor, and its receptor (Nogo-66 receptor, Ng R) were significantly higher than those in the normal control at the acute stage (P<0.01). However, after the intervention of ZGP and YGP, the pathological changes and axon damage in rats' brain and spinal cord were much more alleviated, and the NGF expression was significantly higher than that in the model group at the acute stage (P<0.05). The expression of NGF was even stronger during the remission stage, and a better effect was shown by YGP. As for Nogo A and Ng R expressions, they were significantly lower than those in the model group at the acute stage (P<0.05), but a better effect was shown by ZGP.</p><p><b>CONCLUSIONS</b>ZGP and YGP can prevent axonal injury and promote the axonal regeneration in rats of EAE, and the possible mechanism is to increase the expression of NGF and reduce the expression of Nogo A and its receptor. However, some differences are observed between the two Chinese preparations in their acting times and points, which provides a certain basis for revealing the modern connotation of the Chinese medicine theory on tonifying Shen ()-yin and Shen-yang.</p>
Sujets)
Animaux , Mâle , Rats , Axones , Métabolisme , Anatomopathologie , Physiologie , Encéphale , Métabolisme , Anatomopathologie , Modèles animaux de maladie humaine , Évaluation préclinique de médicament , Médicaments issus de plantes chinoises , Pharmacologie , Encéphalomyélite auto-immune expérimentale , Traitement médicamenteux , Métabolisme , Anatomopathologie , Protéines liées au GPI , Protéines de la myéline , Métabolisme , Facteur de croissance nerveuse , Métabolisme , Régénération nerveuse , Protéines Nogo , Récepteur Nogo-1 , Rats de lignée LEW , Récepteurs de surface cellulaire , Récepteurs peptidiques , Métabolisme , Recherche , Transduction du signal , ComprimésRésumé
<p><b>OBJECTIVE</b>Nogo-A antibody IN-1 can neutralize Nogo-A, a neurite growth inhibitory protein, promoting axonal regeneration following lesions of the central nervous system (CNS) in adult rats. This study aimed to examine the effect of ventricle injection of Nogo-A antibody on neuronal regeneration in neonatal rats following hypoxic-ischemic brain damage (HIBD).</p><p><b>METHODS</b>A model of neonatal HIBD was prepared by the ligation of the left common carotid artery, followed by 8% hypoxia exposure. Forty HIBD rats were randomly given a ventricle injection of 10 microL Nogo-A antibody IN-1 (IN-1 group) or 10 microL artificial cerebrospinal fluid (artificial CSF group) (n=20 each). Another 20 neonatal rats were sham-operated, without hypoxia-ischemia, and were used as the controls. The levels of Nogo-A and GAP-43 protein in the brain were measured by immunohistochemistry.</p><p><b>RESULTS</b>The number of immunohistory positive cells of Nogo-A in the brain in the IN-1 group (28.61+/-1.70) was obviously less than that in the artificial CSF (39.52 +/-1.40) and the sham-operated groups (32.78 +/- 1.87) (both P < 0.01). There were significant differences in the Nogo-A protein expression between the artificial CSF and the sham-operated groups (P < 0.01). The GAP-43 protein expression in the IN-1 group (31.14 +/- 1.88) was noticeably higher than that in the artificial CSF group (27.73 +/- 1.43 ) (P < 0.01). Both the IN-1 and the artificial CSF groups showed lower GAP-43 protein levels than the sham-operated groups (33.64 +/- 1.24) (both P < 0.01).</p><p><b>CONCLUSIONS</b>Nogo-A antibody can reduce the expression of Nogo-A protein in the brain and thus promote neuronal regeneration in neonatal rats following HIBD. An increased GAP-43 protein expression in the brain after Nogo-A antibody administration shows an enhanced neuronal regeneration in the neonatal rats following HIBD.</p>
Sujets)
Animaux , Femelle , Mâle , Rats , Animaux nouveau-nés , Anticorps , Chimie du cerveau , Protéine GAP-43 , Hypoxie-ischémie du cerveau , Métabolisme , Thérapeutique , Immunohistochimie , Injections ventriculaires , Protéines de la myéline , Allergie et immunologie , Régénération nerveuse , Protéines Nogo , Rat Sprague-DawleyRésumé
<p><b>OBJECTIVE</b>Nogo-A is an axon regeneration inhibitor, and its function in central nervous system (CNS) is still unknown. The present study is to explore the relationship between the expression of Nogo-A and the malignancy of oligodendroglial tumors in patients.</p><p><b>METHODS</b>Tumor tissue samples with different malignancy grade were obtained from the hospitals. The samples used for detection had been diagnosed as oligodendroglial tumors (oligodendroglioma or anaplastic oligodendroglioma). The expression of Nogo-A was detected by immunohistochemistry and western-blot analysis. The correlation test between the Nogo-A expression and the morphological changes (the percentages of atypical cells and mitotic cells in the tumors) related to the malignancy of tumor tissues was performed.</p><p><b>RESULTS</b>There was significant negative correlation between the Nogo-A expression and the morphological change of tumor tissues according to immunohistochemistry. Western-blot analysis also indicated that the gray value of Nogo-A protein band in the oligodendroglioma group was significantly higher than that in the anaplastic oligodendroglioma group.</p><p><b>CONCLUSION</b>Nogo-A expression was negatively correlated with the malignancy grade of oligodendroglial tumors.</p>
Sujets)
Adulte , Femelle , Humains , Mâle , Adulte d'âge moyen , Marqueurs biologiques tumoraux , Métabolisme , Tumeurs du cerveau , Diagnostic , Métabolisme , Régulation négative , Physiologie , Immunohistochimie , Index mitotique , Protéines de la myéline , Métabolisme , Invasion tumorale , Diagnostic , Protéines Nogo , Oligodendrogliome , Diagnostic , Métabolisme , Valeur prédictive des testsRésumé
<p><b>OBJECTIVE</b>To screen and identify the proteins that interact with connexin 26 (CX26) and to analyze the expressions of these proteins in cochlea so as to explore the proteins that relate to the trafficking, assembly, localizing and gap junction functions of CX26.</p><p><b>METHODS</b>The whole coding region of GJB2 (CX26) gene was amplified from normal human genomic DNA by polymerase chain reaction (PCR) and then directionally subcloned into the vector pGBKT7 plasmid of the Match Maker Ga14 Two-Hybrid System 3 as a target to screen the interactive proteins of CX26 from the human fetal brain cDNA library by the yeast two hybrid technique. The false positive clones were discarded from the preys by repeated yeast two hybrid method between CX26 and everyone of the preys respectively. The DNAs of the insert of the identified positive clone were sequenced and BLAST analyzed against the GenBank. Lastly, the mRNA of the gene encoding the identified protein was analyzed in the murine inner ear by reverse transcription-polymerase chain reaction (RT-PCR).</p><p><b>RESULTS</b>The insert of one positive clone contained 867 bp with the former 525 bp being coding region. The DNA sequence and the open reading frame of the insert were identical to the 525 bp before the stop codes (including the stop codes) and the 238 bp after the stop codes of RTN4 gene which encoded Nogo protein. And the 174 amino acid residues encoded by the insert were those of the C-terminal of Nogo protein: Nogo-A, Nogo-B and Nogo-C. RTN4 mRNA expressed in the murine inner ear was confirmed by RT-PCR method.</p><p><b>CONCLUSION</b>The C-terminal of Nogo protein interacts with CX26. Nogo protein expresses in the inner ear and may take part in the trafficking of CX26 or CX26 gap junction function.</p>
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Animaux , Humains , Souris , Séquence nucléotidique , Connexine-26 , Connexines , Génétique , Métabolisme , Oreille interne , Métabolisme , Expression des gènes , Données de séquences moléculaires , Protéines de la myéline , Génétique , Métabolisme , Protéines Nogo , Liaison aux protéines , ARN messager , Génétique , Métabolisme , RT-PCR , Techniques de double hybrideRésumé
<p><b>BACKGROUND</b>We investigated the expression and role of TN4 in the oncogenesis of human hepatocellular carcinoma (HCC) from Qidong which is a HCC risk area.</p><p><b>METHODS</b>The expression of TN4 in HCC was observed using immunohistochemical staining (IHC). TN4 levels were manipulated in human liver cancer cell SMMC7721, using pcDNA3.1 eukaryotic expression constructs designed to express the complete TN4 cDNA. The biological changes of the cells were observed before and after transfection of TN4 and the change of gene expression was analysed by atlas cDNA expression array.</p><p><b>RESULTS</b>Among 100 pairs of samples of HCC, TN4 down-regulation expression and up-regulation expression positive rate were 81% (81/100), 19% (19/100), respectively (P < 0.01). TN4 protein was mainly localized in cytoplasm and membrane. The positive rate of TN4 were 10% (3/30), 100% (70/70) in lymph node metastasis and no lymph node metastasis, respectively (P < 0.01). The growth rates of the derivative SMMC7721-TN4 cell lines were decreased in comparison with that of normal SMMC7721 cells and pcDNA-SMMC7721. Some gene expression was changed before and after transfection of TN4. At 30 days of post-implantation of SMMC7721-TN4, SMMC7721-pcDNA3, SMMC7721 group produced tumors of (301.9 +/- 143.4) mm(3), (2418.7 +/- 362.8) mm(3), (2317.4 +/- 587.8) mm(3), respectively, (P < 0.01). Tumor inhibiting rate was 82.4% in TN4 transfection group. Sections of tumors were observed for their degree of tissue necrosis and there was higher degree of necrosis in tumors of the TN4-SMMC7721 cell group than those of the SMMC7721, SMMC7721-pcDNA groups.</p><p><b>CONCLUSIONS</b>TN4 may play an important role in the oncogenesis of human HCC, especially in Qidong, the HCC risk area and TN4 could be a candidate tumor suppressor gene for HCC.</p>