OBJECTIVETo observe the ultrastructural change of the route of gut bacterial translocation in a rat with spinal cord injury (SCI).

METHODSForty Wistar rats were divided into the following groups: control group and 3 SCI groups (10 in each group). The rats in the SCI groups were established SCI model at 24 h, 48 h, and 72 h after SCI. Small intestine mucous membrane tissue was identified and assayed by transmission electron microscope, scanning electron microscope and immunofluorescence microscopy.

RESULTSSmall intestine mucous membrane tissue in control group was not damaged significantly, but those in SCI groups were damaged significantly. Proliferation bacteria in gut lumen attached on microvilli. The extracellular bacteria torn the intestinal barrier and perforated into the small intestinal mucosal epithelial cell. The bacteria and a lot of particles of the seriously damaged region penetrated into the lymphatic system and the blood system directly. Some bacteria were internalized into the goblet cell through the apical granule. Some bacteria and particles perforated into the submucosa of the M cell running the long axis of M cells through the tight junctions. In the microcirculation of mucosa, the bacteria that had already broken through the microvilli into blood circulation swim accompanying with erythrocytes.

CONCLUSIONThe routes of bacterial translocation interact and format a vicious circle. At early step, the transcellular pathway of bacterial translocation is major. Following with the destroyed small intestine mucous, the routes of bacterial translocation through the lymphatic system and the blood system become direct pathways. The goblet cell-dendritic cell and M cell pathway also play an important role in the bacterial translocation.

Animals ; Bacteria ; Bacterial Translocation ; Epithelial Cells ; microbiology ; Goblet Cells ; microbiology ; Intestinal Mucosa ; microbiology ; pathology ; ultrastructure ; Intestine, Small ; microbiology ; pathology ; ultrastructure ; Microvilli ; microbiology ; Rats ; Rats, Wistar ; Spinal Cord Injuries ; microbiology

OBJECTIVETo document ultrastructural changes of brain, spinal cord, skeletal muscle, jejunum and lung of EV71 infection mouse model, and to explore the myotropism and pathogenesis of EV71 in nervous system.

METHODSTen-day-old suckling mice were infected with EV71 strain via the intraperitoneal route. Mice with paralysis were scarified on day 4 post infection and the brain, spinal cord, skeletal muscle, jejunum and lung were sampled for transmission electron microscopy and light microscopy.

RESULTSLesions in brain were generally mild with inner chamber swelling in some of mitochondria. Myelin sheaths of medullated fibers were split with vacuolated changes. The Nissl bodies in anterior motor neurons disappeared along with mitochondria swelling, rough endoplasmic reticulum swelling and degranulation. Cytoplasm of anterior motor neurons showed cribriform appearance accompanied by neuronophagia. The bands of skeletal muscle in the infected group disappeared with degeneration and karyopyknosis in myocytes, in addition to mitochondrial swelling. Microvilli of epithelium in jejunum became loosely arranged along with formation of spiral medullary sheath structure and mitochondria swelling. Interstitial pneumonia was observed in lungs with type II pneumocyte proliferation and evacuation of the multilamellar bodies.

CONCLUSIONSEV71 infection causes severe myositis in the mouse model suggesting a strong myotropism of EV71 virus. The presence of lesions of various degrees in central nervous system and changes in anterior motor neurons may be associated with limb paralysis.

Animals ; Brain ; ultrastructure ; virology ; Disease Models, Animal ; Enterovirus A, Human ; Enterovirus Infections ; pathology ; virology ; Jejunum ; ultrastructure ; virology ; Lung ; ultrastructure ; virology ; Mice ; Mice, Inbred BALB C ; Microscopy, Electron, Transmission ; Muscle, Skeletal ; ultrastructure ; virology ; Spinal Cord ; ultrastructure ; virology

OBJECTIVETo study the role and effect of Schwann cells (SCs) remyelination in contused spinal cord.

METHODSGreen fluorescence protein expressing-SCs were transplanted into the epicenter, rostral and caudal tissues of the injury site at 1 week after the spinal cords were contused. At 6 weeks, the spinal cords were removed for cryosections, semithin sections and ultrathin sections, and then immunocytochemical staining of myelin basic protein (MBP), P0 protein (P0) and S100 protein (S100) was carried out on the cryosections. Qualitative and semiquantitative analyses were performed on the cryosections and semithin sections. Ultrastructure of myelinated fibers was observed on the ultrathin sections under electron microscope.

RESULTSTransplanted SCs and myelinated fibers immunocytochemically labeled by MBP, P0 as well as S100 distributed in whole injured area. The quantity of myelinated fibers labeled by the three myelin proteins showed no statistical difference, however, which was significantly larger than that of controls. On the semithin sections, the experimental group demonstrated more myelinated fibers in the injured area than the controls, but the fibers had smaller diameter and thinner myelin sheath under electron microscope.

CONCLUSIONSCs can promote regeneration of injured nerve fibers and enhance remyelination, which may be histological basis of SCs-mediated functional repair of injured spinal cords.

Animals ; Immunohistochemistry ; Microscopy, Electron ; Myelin Basic Protein ; metabolism ; Myelin P0 Protein ; metabolism ; Nerve Regeneration ; physiology ; Rats ; Rats, Sprague-Dawley ; S100 Proteins ; metabolism ; Schwann Cells ; physiology ; ultrastructure ; Spinal Cord Injuries ; metabolism ; physiopathology

OBJECTIVE: To investigate the behavioral and ultrastructural changes of intrathecal administration of different concentrations of ropivacaine for 12 h. METHODS: Thirty male SD rats were randomly divided into 5 groups (6 rats in each group):group N (control), group A (ropivacaine 0.25%), group B (ropivacaine 0.5%), group C (ropivacaine 0.75%),and group D (ropivacaine 1.0%). A polyurethane microcatheter was inserted into the lumbar subarachnoid space 8 cm according to Yakshos intrathecal administration. The rats in group N received saline 0.12 mL/kg for 8 times at 1.5 h interval through the catheter, and the rats in the other groups received different concentrations of ropivacaine in the same way as in group N. The poster paw withdrawal latency to heat (PWHL) and mechanical stimulation (von Fray filament) (PWML) were measured the day before the intrathecal administration and 12 hours after the first intrathecal administration of ropivacaine. Motor function (MF) was measured after the last intrathecal administration. After the behavior test, the rats were sacrificed and the lumber segments of the spinal cord were immediately removed for electron microscopic examination. RESULTS: A total hind limb paralysis was seen at 30 seconds and intramuscular strain gradually came back 10~60 minutes after the intrathecal administration of ropivacaine in group A, B, C, and D, but not in group N. The recovery time of motor block of group A was the shortest (P<0.05), that of group D was the longest,and that of group B and C was between group A and D. Intrathecal administration of different concentrations of ropivacaine did not affect the percent maximum possible effect (%MPE) of PWHL and PWML. Electron microscopic examination showed that the spinal cords were normal in group N and A, slight edema of mitochondria and endoplasmic reticulum (ER) in group B, loosened fibrous layers in medullary sheath, edema and local degeneration of neuraxis in group C,and shrinkage of nuclear membrane, serious edema of ER, vacuolus change of mitochondria and local demyelination in group D. CONCLUSION Ropivacaine (0.5%, 0.75%, and 1.0%) administered intrathecally for 12 hours causes different degrees of ultrastructural changes in the spinal cord depending on concentrations.
Amides ; administration & dosage ; toxicity ; Anesthetics, Local ; administration & dosage ; toxicity ; Animals ; Behavior, Animal ; drug effects ; Dose-Response Relationship, Drug ; Injections, Spinal ; Male ; Microscopy, Electron ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Ropivacaine ; Spinal Cord ; ultrastructure

OBJECTIVECombine olfactory ensheathing glia (OEG) implantation with ex vivo non-viral vector-based neurotrophin-3 (NT-3) gene therapy in attempting to enhance regeneration after thoracic spinal cord injury (SCI).

METHODSPrimary OEG were transfected with cationic liposome-mediated recombinant plasmid pcDNA3.1(+)-NT3 and subsequently implanted into adult Wistar rats directly after the thoracic spinal cord (T9) contusion by the New York University impactor. The animals in 3 different groups received 4x10(5) OEG transfected with pcDNA3.1(+)-NT3 or pcDNA3.1(+) plasmids, or the OEGs without any plasmid transfection, respectively; the fourth group was untreated group, in which no OEG was implanted.

RESULTSNT-3 production was seen increased both ex vivo and in vivo in pcDNA3.1(+)-NT3 transfected OEGs. Three months after implantation of NT-3-transfected OEGs, behavioral analysis revealed that the hindlimb function of SCI rats was improved. All spinal cords were filled with regenerated neurofilament-positive axons. Retrograde tracing revealed enhanced regenerative axonal sprouting.

CONCLUSIONNon-viral vector-mediated genetic engineering of OEG was safe and more effective in producing NT-3 and promoting axonal outgrowth followed by enhancing SCI recovery in rats.

Animals ; Animals, Newborn ; Brain Tissue Transplantation ; methods ; Cells, Cultured ; DNA, Recombinant ; therapeutic use ; Disease Models, Animal ; Female ; Gene Transfer Techniques ; Genetic Therapy ; methods ; Genetic Vectors ; genetics ; Graft Survival ; genetics ; Growth Cones ; metabolism ; ultrastructure ; Nerve Regeneration ; genetics ; Neuroglia ; metabolism ; transplantation ; Neurotrophin 3 ; biosynthesis ; genetics ; Olfactory Bulb ; cytology ; transplantation ; Paralysis ; metabolism ; physiopathology ; therapy ; Plasmids ; genetics ; Rats ; Rats, Wistar ; Recovery of Function ; genetics ; Spinal Cord Injuries ; metabolism ; physiopathology ; therapy ; Treatment Outcome ; Up-Regulation ; genetics

BACKGROUNDLittle is known about neuronal death mechanisms following spinal cord ischemia. The present study aimed to investigate the protective effect of pentoxifylline (PTX) against spinal cord ischemia/reperfusion (I/R) injury.

METHODSRabbits sustained spinal cord ischemia following 45 minutes cross-clamping of the infrarenal aorta. Experimental groups were as follows: the first group of animals (sham, n = 8) underwent laparotomy alone and served as the sham group; the second group (I/R, n = 20) received carrier (3 ml saline solution) and served as the control group; the third group (PTX-A, n = 20) received PTX intravenously 10 minutes prior to ischemia; and the fourth group (PTX-B, n = 20) received PTX intravenously at the onset of reperfusion. Rabbits were evaluated for hind-limb motor function with the Tarlov scoring system at 48 hours. Serum was assayed with enzyme-linked immunosorbent assay for tumor necrosis factor alpha (TNF-alpha) and spinal cords were harvested for myeloperoxidase (MPO) activity, histopathological analysis, terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling staining, platelet/endothelial cell adhesion molecule-1 (PECAM-1) and caspase-3 immunohistochemistry, and the number of necrotic and apoptotic neuron were counted and data analyzed at 12, 24, 48 and 72 hours of reperfusion. Spinal cords were studied by electron microscopy.

RESULTSImproved Tarlov scores were seen in PTX-treated rabbits as compared with ischemic control rabbits at 48 hours. A significant reduction was found in TNF-alpha in serum, activity of MPO and immunoreactivity of the PECAM-1 and caspase-3 in PTX-treated rabbits. There were fewer apoptotic neurons than necrotic neurons (P < 0.05). A significant decrease in both necrotic and apoptotic neurons was observed in the PTX-treated groups (PTX-A and PTX-B) compared with the I/R group (P < 0.05). Both necrotic and apoptotic neurons were found with the electron microscope.

CONCLUSIONSPTX may induce protection against ischemia injury in the spinal cord, thereby preventing both necrosis and apoptosis. A major mode of cell death in spinal cord ischemia/reperfusion injury is necrosis while apoptosis is not dominant.

Animals ; Apoptosis ; drug effects ; Caspase 3 ; metabolism ; Immunohistochemistry ; In Situ Nick-End Labeling ; Microscopy, Electron, Transmission ; Necrosis ; Pentoxifylline ; pharmacology ; therapeutic use ; Rabbits ; Reperfusion Injury ; prevention & control ; Spinal Cord ; blood supply ; pathology ; ultrastructure ; Spinal Cord Ischemia ; prevention & control ; Vasodilator Agents ; pharmacology ; therapeutic use

OBJECTIVE: To explore the alterations of serum of myelin basic protein (MBP) concentration in the plasma and ultrastructure in the spinal cord after continuous intrathecal injection of different ropivacaine concentrations in rats. METHODS: Ninety-six male Sprague-Dawley rats weighing 220 to approximately 280 g were randomly divided into a control group (Group N), Group R(1), R(2) and R(3) (24 rats in each group). Each group was subdivided into 4 subgroups (6 rats in each subgroup). According to the method of Yaksh's, a polyurethane microspinal catheter was inserted into the lumbar subarachnoid space in which 8 cm segment was left. Rats in each group were continuously received 40 microL of intrathecal injection of normal saline(Group N), 0.5%, 0.75%, and 1.0% ropivacaine (Group R(1),R(2),R(3)), 3 times every 1.5 hours. Blood (0.5 mL) was drawn from the femoral artery to determine serum concentrations of MBP at the detecting time T(0)(before inserted pipe)and T(1)(before the first intrathecal injection); for the subgroups, the examining time was at T(2), T(3), T(4) and T(5)(6, 12, 24 and 48 h respectively after the last time intrathecal administration). After blood was drawn, the rats in each subgroups were decapitated and the spinal cord of L(1-2) intumescentia lumbalis were immediately removed for electronic microscopic examination. RESULTS: MBP levels were comparatively steady in Group N, R(1) and R(2), while there was statistical difference between Group R(3) and Group N, R(1),R(2),and R(3) (P<0.05). MBP level of Group R(3) was significantly higher at T(2),T(3),T(4) and T(5) than that at T(0)(P<0.01). The ultrastructural changes of the spinal cord in Group R(3) were pycnosis of most neurons, dilation of most rough endoplasmic reticulum, and vague structure of mitochondria and endocytoplasmic reticulum. A few neurons were completely de-generated losing the normal structure, with vacuole degeneration of crista mitochondriales or even partial loss. CONCLUSION The spinal cord ultrastructure is selectively vulnerable after intrathecal 1.0% ropivacaine injection, which may be one of the important pathophysiological bases for local anesthetic neurotoxicity. MBP may serve as a sensitive and specific indicator of spinal cord damage after intra-thecal administration of ropivacaine.
Amides ; administration & dosage ; pharmacology ; Anesthetics, Local ; administration & dosage ; pharmacology ; Animals ; Injections, Spinal ; methods ; Male ; Myelin Basic Protein ; Nerve Tissue Proteins ; blood ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Ropivacaine ; Spinal Cord ; ultrastructure ; Transcription Factors ; blood

OBJECTIVETo investigate the recovery of rat transected spinal cord injury after implantation of Schwann cells combined with poly (lactide-co-glycolide) (PLGA).

METHODSSchwann cells were expanded, co-cultured with PLGA for 9 days in vitro, and then analyzed with scanning electron microscope (SEM). Rat spinal cord at the level of T(9) was transected. Schwann cells labeled with BrdU and PLGA scaffold were implanted to injury site. After 1, 3, 6 months, BrdU/MBP immunohistochemistry double staining, semi-thin sections stained thionin and ultra-thin section were performed to investigate myelin renew. BBB open field locomotion, motor evoked potential (MEP), compound muscle action potential (CMAP) and somatosensory evoked potential (SEP) were recorded.

RESULTSSchwann cells grew well on PLGA under SEM. BrdU/MBP double positive cells would been seen, remyelination was thin and formed by Schwann cells at 6 months later under electron microscope (EM). BBB behavioral tests revealed no significant difference in recovery comparing with experiment group and control group. The results of MEP, CMAP and SEP showed no significant improvement in the conduction of spinal cord.

CONCLUSIONSThere are the compatibility between Schwann cells and PLGA. Although remyelination was found in morphology, function conduction of spinal cord failed to be established.

Animals ; Cells, Cultured ; Disease Models, Animal ; Evoked Potentials, Motor ; Female ; Immunohistochemistry ; Lactic Acid ; chemistry ; Microscopy, Electron, Scanning ; Microscopy, Electron, Transmission ; Nerve Regeneration ; Polyglycolic Acid ; chemistry ; Prostheses and Implants ; Rats ; Rats, Wistar ; Schwann Cells ; chemistry ; transplantation ; ultrastructure ; Spinal Cord Injuries ; physiopathology ; surgery ; Tissue Engineering ; methods

Clear cell ependymoma was included in the World Health Organization classification of the nervous system in 1993, and all the reported cases, except for two in the spinal cord, were located in the brain, mainly in the supratentorial compartment. Astrocytomas outnumber ependymomas in the spinal cord, and the two entities partly share cytologic findings such as long, bipolar glial processes and oval to round nuclei resembling those seen in pilocytic astrocytoma. Here, we report the first Korean case of intramedullary clear cell ependymoma of the spinal cord, which is the third case situated in the spinal cord in the literature. The crush smear revealed round-to-oval nuclei with occasional nuclear eosinophilic inclusion and rare nuclear grooves. Cytoplasm had fluffy eosinophilic glial processes, and acellular fibrillary zone. On hematoxylin-eosin stain, oval to round tumor cells had large central nuclei with indistinct nucleoli and a moderate amount of clear cytoplasm, i.e. perinuclear halo, mimicking oligodendroglioma. Perivascular pseudorosettes and ependymal clefts were rarely found. In retrospect, perinuclear halo was absent on crush smears. Ultrastructurally, they had extensive surface microvilli and edematous cytoplasm filled with abundant glial filaments and microlumens with or without microvilli. Intercellular long cell junctions of the zipper-like zonula adherens type were found.
Aged ; Ependymoma/metabolism/*pathology/ultrastructure ; Female ; Humans ; Immunohistochemistry ; Microscopy, Electron, Transmission ; Spinal Cord Neoplasms/metabolism/*pathology/ultrastructure

By using stereological morphometric techniques, we examined the ultrastructure of synapses in lamine II of the spinal dorsal horn of Sprague Dawley rats 30 min, 3 h and 5 h after long-term potentiation (LTP) induction. We found that the numerical density per unit volume (Nv) of total synapses, the thickness of the postsynaptic density (PSD), width of the synaptic cleft increased significantly after the establishment of LTP. (1) Thirty minutes after the formation of LTP, the thickness of the PSD increased from 0.029 +/-0.0064 microm (control) to 0.036 +/-0.009 microm (P<0.05) and the width of the synaptic cleft increased from 0.0181+/-0.0024 microm (control) to 0.0197+/-0.0029 microm (P< 0.05); the number of synaptic vesicles decreased from 0.122 +/-0.011/microm(2) to 0.085 +/-0.010/microm(2) (P<0.05); (2) 3 h after the formation of LTP, the thickness of PSD and the width of the synaptic cleft had no difference compared with those 30 min after LTP. The number of synaptic vesicles increased from 0.122 +/-0.011/microm(2) to 0.138 +/-0.015/microm(2); the curvature of the synaptic interface increased from 1.153+/-0.195 to 1.386 +/-0.311 (P<0.05, compared with control). Nv of negative synapses increased from 0.0187 +/-0.0056 to 0.0543 +/-0.0152 (P<0.05, compared with control), Nv of perforated synapses also increased from 0.0135 +/-0.0053 to 0.0215 +/-0.0076 (P<0.05, compared with control). These data suggest that the increase in thickness of PSD might be the major morphological change during the induction of LTP, while the increase in curvature of the synaptic interface, and the number of perforated synapses might be responsible for the maintenance of the spinal LTP.
Animals ; Long-Term Potentiation ; physiology ; Male ; Posterior Horn Cells ; physiology ; ultrastructure ; Rats ; Spinal Cord ; anatomy & histology ; physiology ; Synapses ; ultrastructure ; Synaptic Transmission ; Synaptic Vesicles ; ultrastructure