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Human central nervous system primitive neuroectodermal tumor expressing nerve growth factor receptors: CHP707m.

A primitive neuroectodermal tumor (PNET) presented as a cerebral hemispheric mass in a 33-year-old man. Bone marrow metastases were discovered 11 months later. A cell line (CHP707m) was derived from these metastases. In culture, the cells showed features of neuronal differentiation, forming short neurites and synthesizing low-molecular-weight neurofilament protein. Northern blotting showed the tumor cells express nerve growth factor (NGF) receptor messenger RNA, and fluorescence-activated cell-sorting demonstrated NGF receptors on the cell surface. Western blotting showed CHP707m NGF receptors are truncated. The receptors are functional; they bind iodine 125-labeled mouse NGF with an affinity of 1.6 x 10(-9) M, and short-term treatment with NGF induces expression by the tumor cells of the proto-oncogene, c-fos. Although CHP707m is the first central nervous system PNET cell line proven to express NGF receptors, immunohistological survey of tissue sections prepared from human central nervous system PNETs showed that 13 of 35 contained NGF receptor-positive tumor cells. Thus, more than one-third of such tumors might be responsive to the effects of NGF.

Adult↗

Quantification of hoof deformation using optical motion capture.

Deformation of the hoof capsule occurs during stance phase as a result of loading by the distal phalanx. The objective of this study was to evaluate optical motion capture as a technique for measuring hoof deformation during locomotion, and to quantify the deformation of the dorsal wall during stance phase. The front left feet of 3 clinically sound horses were used for this study. The hoof wall was cleaned and 9 retroreflective markers were glued to the lateral and dorsal hoof wall, and shoe. Each horse was trotted in a straight line at constant speed while marker positions during stance phase were recorded at 240 frames/s using a 3-dimensional (3-D) motion capture system. Similar patterns of hoof deformation were seen in all horses; the dorsal wall moved inwards towards the axis of the limb with displacement increasing proximally. Mean peak deformation varied between 0.17 and 0.92 mm in the dorsal hoof wall, and 0.18 and 0.76 mm in the lateral hoof wall. In all horses, movement of the shoe relative to the ground was < 0.7 mm throughout stance phase and movement of the hoof relative to the shoe was < 0.3 mm. Calibration using a micrometer indicated the motion capture system was accurate to < 0.1 mm in the configuration used for the study. This study showed that optical motion capture can be used to measure hoof deformation and successfully addresses some of the limitations of existing methods.

Animals↗

Synchronous measurements of blood pressure and red blood cell velocity in capillaries of human skin.

We have devised a system for analyzing the temporal coherence of capillary pressure (CP) and capillary red blood cell velocity (CBV) variations in humans. The system is designed to measure human blood pressure in skin capillaries by direct cannulation while simultaneously measuring red blood cell velocity in the same capillary by video microscopy. The servo-nulling pressure measurement system allows the dynamic recording of capillary pressure with a flat frequency response of about 12 Hz. Computerized data acquisition is synchronized with the frame code of a U-matic video recorder, which records the capillaroscopic picture for later computerized off-line analysis of capillary red blood cell velocity. Measurements of simultaneously recorded CP and CBV in healthy volunteers show synchronized pulsation. Minimal pulsatile variations in CP values result in marked variations of CBV. In addition, minor fluctuations (3-4 cycles per minute) in CP are accompanied by marked changes in CBV. This system may help provide information about microvascular pathophysiology in skin diseases before and after treatment.

Adult↗

Computer-assisted measurement of the size of ossification in patients with ossification of the posterior longitudinal ligament in the cervical spine.

BACKGROUND: Progression of ossification of the posterior longitudinal ligament in patients may lead to serious neurological deterioration. A government-funded study group established a manual method of measurement on plain radiographs to detect progression of the ossified lesion. However, this method did not gain wide acceptance because it was time-consuming and complicated, for which drawings of many lines and points are required. We have applied a computer-assisted measurement system to this task and have evaluated inter- and intraexaminer reliability, showing that it is quicker to use and more accurate than the manual method. METHODS: Eight board-certified spine surgeons, acting as the examiners, measured the sizes of the ossified lesions on nine lateral cervical spine radiographs using the computer-assisted measurement system. Following insertion of digitized radiographic image data into a computer, the corners of the vertebral bodies on the displayed images are marked by the examiners, and the software automatically sets reference lines and points. The examiners identify upper, lower, and posterior margins of the ossified lesions, and the software calculates the dimensions of the ossified lesions. Data obtained from eight examiners for length and thickness underwent rigorous statistical analysis by calculating the intraclass correlation coefficients with 95% confidence intervals (CIs) to determine interexaminer reliability and Pearson's correlation coefficients between the two measurements by the same examiner to determine intraexaminer reliability. RESULTS: The intraclass correlation coefficients were 0.927 and 0.968 with 95% CIs of 0.883-0.955 and 0.956-0.978 for measurements of length and thickness, respectively, of the ossified lesions. The Pearson's correlation coefficients for the two measurements by the same examiners were 0.943-0.985 for length and 0.957-0.991 for thickness. CONCLUSIONS: The inter- and intraexaminer reliability using this measurement system was excellent. The method can detect progression of ossification of the posterior longitudinal ligament (OPLL) on plain radiographs with high precision and could become a standard method for measuring the size of OPLL.

Cervical Vertebrae↗

Central inhibitory interactions in human vision.

Contrast threshold and perceived orientation of a line segment were measured when another line segment was simultaneously presented either to the same or the other eye; the angle between the two line segments was varied. The presence of the masking line elevated the contrast threshold under both conditions and the threshold increased similarly both in monoptic and dichoptic masking when the masking angle was made smaller. The presence of the masking line affected also the perceived orientation of the test line. The effect was similar both in monoptic and dichoptic masking; the largest change of perceived orientation occurred at about 15 degrees masking angle and the change was smaller at other angles. The effect disappeared within a short distance when the masking line was removed farther from the test line. The similarity of the monoptic and dichoptic threshold elevations demonstrates that there are lateral inhibitory interactions between central neural units in the human visual system. It is likely that the interacting units mediate the perception of contour orientation, for the threshold elevation functions were consistent with concurrent changes of perceived orientation. The results are evidence for the hypothesis that inhibition between orientation detectors is a factor in the perceptual expansion of acute angles.

Humans↗

Computer programs for the radioactive microsphere technique. Determination of regional blood flows and other haemodynamic variables in different experimental circumstances.

Computer programs in FORTRAN IV have been designed to calculate regional blood flow values from injections of gamma radiation-emitting radioactive microspheres. The first program PRSO constitutes a file containing the names of organs, organ groups and systemic haemodynamic variables and data about the isotopes in use. The second program PRSI transfers the data of the particular experiment concerning protocol, systemic haemodynamic variables, organ weights, number of vials belonging to each organ and the radioactivity from each vial into a data file that is subsequently analyzed by the third program PRSII. One of 5 data conditions which later directs calculations in PRSII is determined by PRSI. PRSII deducts background radioactivity, decomposes the spectrum from the mixture of up to 6 nuclides using the "stripping' method, and corrects for isotope decay during the counting period. Depending upon the data condition, PRSII calculates one or more of the following variables: (i) cardiac output (CO); (ii) the distribution of CO; (iii) regional blood flow values and tissue vascular resistances; (iv) the distribution of blood flow within an arterial bed; (v) arteriovenous anastomotic blood flow; (vi) some specified systemic haemodynamic variables; and (vii) blood flow ratios. The systemic and regional haemodynamic data belonging to experiments are saved for later statistical analyses (e.g., mean, standard error of the variables, changes from the base line, comparison between two groups of experiments) by other programs. PRSO, PRSI and PRSII can be operated by investigators with little or no special training in computer use.

Animals↗

An inability of gamma-acetylenic GABA to block eating evoked by hypothalamic stimulation.

Several lines of evidence have implicated the nigrostriatal dopamine system and the neocortex for eating occurring either spontaneously or in response to electrical stimulation of the lateral hypothalamus (ESLH). GABA-ergic neural systems are known to modulate activity in the nigrostriatal system and neocortex. The GABA-transaminase inhibitor, gamma-acetylenic GABA (GAG) was administered to rats eating in response to ESLH. This drug produced hypersynchronization of the cortical EEG and behavioral sedation, but did not alter the current threshold for evoking eating by ESLH. Implications of results for understanding mechanisms underlying ESLH-induced eating are discussed.

Alkynes↗

Chemokine signaling: rules of attraction.

The chemokine SDF-1 and its receptor CXCR4 control cell migration in the immune and nervous systems. Recent studies in zebrafish have shown that SDF-1 and CXCR4 also guide the migration of germ cells and sensory organs of the lateral line.

Animals↗

Comparative pharmacophore modeling of organic anion transporting polypeptides: a meta-analysis of rat Oatp1a1 and human OATP1B1.

The organic anion transporting polypeptides OATPs are key membrane transporters for which crystal structures are not currently available. They transport a diverse array of xenobiotics and are expressed at the interface of hepatocytes, renal tubular cells, enterocytes, and the choroid plexus. To aid the understanding of the key molecular features for substrate-transporter interactions, pharmacophore models were produced for the two OATPs that have been most extensively studied, namely rat Oatp1a1 and human OATP1B1. Literature data from Chinese hamster ovary, HeLa, human embryonic kidney 293 cells, and Xenopus laevis oocytes were used to construct pharmacophores for each individual transporter which were later merged to show similarities across cell lines for the same transporter. Additionally, meta-pharmacophores were generated from the combined datasets of each cell system used with the same transporter. The pharmacophores for each transporter consisted of hydrogen bond acceptor and hydrophobic features. There was good agreement between the merged and meta-pharmacophores containing two hydrogen bond acceptors and two or three hydrophobic features for Oatp1a1 and OATP1B1. External test sets were used to validate the individual pharmacophores. The meta-pharmacophores were also used to make predictions for molecules not included in the models and provided new molecular insight into the key features for these OATP transporters. This approach can be extended to other transporters for which limited data are available.

Animals↗

Overproduction of Rb protein after the G1/S boundary causes G2 arrest.

The Rb protein is known to exert its activity at decision points in the G1 phase of the cell cycle. To investigate whether it may also play some role(s) at later points in the cell cycle, we used a system of rapid inducible gene amplification to conditionally overexpress Rb protein during G2 phase. A cell line expressing a temperature-sensitive simian virus 40 large T antigen (T-Ag) was stably transfected with plasmids containing the Rb cDNA linked to the simian virus 40 origin of replication: pRB-wt, pRB-fs, and pRB-Dra, carrying wild-type murine Rb cDNA, a frameshift mutation close to the beginning of the Rb coding region, and a single-amino-acid deletion in the E1A/T-Ag binding pocket, respectively. Numerous independent cell lines were isolated at the nonpermissive temperature; cell lines displaying a high level of episomal amplification of an intact Rb expression cassette following shiftdown to the permissive temperature were chosen for further analysis. Plasmid pRB-fs did not express detectable Rb antigen, while pRB-Dra expressed full-length Rb protein. The Dra mutation has previously been shown to abrogate phosphorylation as well as T-Ag binding. Fluorescence-activated cell sorting (FACS) analysis revealed that cultures induced to overexpress either wild-type or Dra mutant Rb proteins were significantly enriched for cells with a G2 DNA content. Cultures that amplified pRB-fs or rearranged pRB-wt and did not express Rb protein had normal cell cycle profiles. Double-label FACS analysis showed that cells overexpressing Rb or Rb-Dra proteins were uniformly accumulating in G2, whereas cells expressing endogenous levels of Rb were found throughout the cell cycle. These results indicate that Rb protein is interacting with some component(s) of the cell cycle-regulatory machinery during G2 phase.

Animals↗

Characterization and evaluation of detoxification functions of a nontumorigenic immortalized porcine hepatocyte cell line (HepLiu).

Primary porcine hepatocytes (PPH) are currently used in research and therapeutic applications as the biological component of extracorporeal liver assist devices to overcome the shortage of human hepatocytes. However, their finite life span and typically rapid loss of functions limit their utility. An immortalized, nontumorigenic, highly differentiated porcine hepatocyte cell line was developed in our laboratory to resolve these disadvantages. PPH were transfected with simian virus 40 (SV40) T antigen under the control of the SV40 early promoter. From the established 69 clones, 23 clones displaying hepatocyte-like morphology were screened for diazepam metabolism. One clone, HepLiu D63, has been maintained in culture for > 2 years, through more than 60 passages and 240 divisions. Albumin protein, present in early passages, was lost at later passages, but albumin transcript still was detectable in later passages. Carbamoyl phosphate synthetase, a gateway enzyme of the urea cycle, was consistently detectable in HepLiu cells. Cytokeratin 18, a characteristic marker of primary hepatocytes, was detected by both immunofluorescent staining and Western blot in HepLiu cells. Furthermore, maintenance of P450 functions in HepLiu cells was evidenced by diazepam and 7-ethoxycoumarin metabolites measured by HPLC. Phase II conjugative function was measured as acetaminophen glucuronidation. P450 dealkylase was demonstrated microscopically by the conversion of a nonfluorescent substrate to a fluorescent product. Both Northern blot analysis and immunofluorescent staining showed SV40 T antigen expression in the nuclei of HepLiu cells. No tumor formation occurred when HepLiu cells were injected into severe combined immunodeficient (SCID) mice nor was the TAI (a tumor marker) mRNA expressed, even in later passages. This immortalized, nontumorigenic, highly functional cell line may provide a valuable tool for drug/toxicological studies, liver biologic regulation studies, and artificial liver support systems.

Animals↗

[Frequency characteristics of central neurons of the electroreceptor system of the skate in response to exposure to electric and magnetic fields].

Responses of single units to sinusoidal homogenous electrical and magnetic field stimulation were studied in the lateral-line lobes of the skate Raja radiata. The voltage gradient thresholds were in the range of 0.03-10 microV/cm. Optimal frequency ranges for electrical and magnetic stimuli were 0.05-0.5 and 2-3 Hz, respectively. Possible mechanisms and functional significance of frequency characteristics of the neurons are discussed.

Animals↗

Electron microscopic examination of lead treated L6 skeletal muscle line cells in culture.

A study has been made of the effects of lead on developing muscle using a cell culture system. When the L6 clonal cell line was treated with lead acetate in culture during the first or second day after plating (but not at later times) fusion of cells to form skeletal muscle straps was inhibited. This inhibition was not due to a general toxic effect of lead on the cells since cell division continued at a normal rate during the first 10 days of culture after lead addition. When cells were examined ultrastructurally using stereology, the only apparent effect of lead was a change in mitochondrial size and configuration. Five times as many partially condensed mitochondria were seen in lead treated cells, and mitochondrial size was also significantly increased. Lead appears to effect a very early event in cell fusion since gap junction formation (a necessary event in fusion) and some synthesis of myofibrils occur even in the presence of lead.

Cell Fusion↗

Human fetal heart development after mid-term: morphometry and ultrastructural study.

A total of 44 normally developed human fetal hearts aged from 17 to 40 weeks gestation were provided for routine ultrastructural and morphometric studies. For morphometric analysis, the maximal thicknesses of the anterior, lateral and posterior wall of both ventricles and that of interventricular septum were measured and the left-to-right thickness ratios calculated. The cross-sectional areas of both ventricles in the plane of the greatest heart diameter were measured with computerized image analysis system. Data were analyzed using linear regression and one-way analysis of variance. Myofibril formation occurred by attachment of thin filaments into amorphous Z materials which were presented in sarcolemmal plaques, sarcoplasmic condensations, desmosomes and in Z lines. From these Z centers, myofibrils radiated many directions and branched and anastomosed with further development. This pattern of myofibrillar development continued throughout the whole fetal period. A transverse tubule system was clearly evident in later fetal development. It occurred by invagination of sarcolemma into myocardial cells and formation of subsarcolemmal caveolae. Mitochondria, well-developed Golgi complexes, glycogen granules and well-developed microvessels were found throughout the whole fetal period. Binucleated myocytes appeared by 32 weeks gestation and this suggests that myocyte hyperplasia may cease before birth in humans. The growth of both ventricular walls, the interventricular septum and that of both ventricular cross-sectional areas showed linear regression, and the left-to-right wall thickness ratios were nearly constant. Also, there were no differences in morphometric data between the left and right ventricles. In conclusion, development of the myocyte is an ongoing process which may be continued in the post-natal period in humans, and our statistical results do not support the theory of the right ventricular dominance during the fetal period.

Cell Differentiation↗

In vivo adenoviral transduction of the neonatal rat cochlea and middle ear.

Virally mediated gene transfer to the adult mammalian ear appears to be a powerful strategy to investigate gene function in the auditory system and to develop new therapeutic treatment for hearing impaired patients. However, there has been little work done in the neonatal middle and inner ear. In this study, a recombinant adenoviral (AdV) vector was used for gene transfer of a beta-galactosidase (beta-gal) reporter gene to the neonatal middle ear and cochlea of 5 day old rats. For transduction of middle ear, AdV was injected through the tympanic membrane into the tympanic cavity. Three and 7 days later, strong expression of beta-gal was observed in epithelial cells of the mucosa, but not in the underlying stroma or mesenchyme. There was little or no infiltration of leukocytes. No expression of beta-gal was detected inside the cochlea or vestibular system. When AdV was injected into the basal turn of the cochlea, high levels of beta-gal expression were observed in cells lining the perilymphatic space and in parts of the spiral ligament 3, 7 and 21 days later. Spiral ganglion cells did not express beta-gal. However, virally mediated gene transfer was observed in some cells of the organ of Corti. A moderate infiltration of leukocytes into the labyrinth was observed, but no vestibular or auditory dysfunction. These results demonstrate that neonatal middle ear and cochlear cells can be successfully transduced with an AdV vector in vivo, without obvious morphological signs of inflammation or cellular damage. AdV vectors provide a tool for investigation of the role of genes in influencing the development of middle and inner ear structures. Virally mediated expression of protective genes could also be used to rescue hair cells or spiral ganglion cells from congenital degeneration or damage.

Adenoviridae↗

Distribution of glutamate decarboxylase-like immunoreactivity in the sixth abdominal ganglion of the cockroach Periplaneta americana.

An antiserum against glutamate decarboxylase (GAD) of the rat brain was used to locate GAD activity in sections of the nervous system of the cockroach, Periplaneta americana. The sixth abdominal ganglion was chosen because electrophysiological evidence suggests the presence of GABAergic inhibitory synapses in the cercal-giant interneuron system. Groups of somata and numerous fibres and tracts were positively labelled by the GAD antiserum. A posterior group of labelled somata could be identified close to the entry of the cercal nerves. A line of somata clusters lay along a ventro-lateral furrow. Another discrete row of GAD-like cells was located dorso-laterally. Some small cells among the dorsal unpaired neurons were labelled. A small central group appeared under these cells. An abundance of GAD-like processes and transversal tracts were found within the neuropile. The different systems of GABAergic inhibitors in the ganglion are discussed; in particular we show that the fibres of cercal nerve X are not labelled. This demonstrates that the latter act on the giant fibres via interneurons. We suggest that the group that sends axons into the overlapping region between the cercal nerve and the giant fibre could be the inhibitory interneurons involved in this system.

Abdomen↗

Perioperative outcome of laparoscopic left lateral liver resection is improved by using staple line reinforcement technique: a case report.

Current laparoscopic stapling technology still cannot prevent bile leakage (<55%) along the resection margin. A new staple line reinforcement technique was used in the present case. This technique incorporates an absorbable polymer membrane into the stapler system, such that it buttresses the transected solid organ. The objectives of this novel procedure are to decrease hemorrhage at the staple line and to prevent bile duct leakage after liver resection. A 47-year-old man followed for status post biliopancreatic diversion with duodenal switch presented with epigastric pain. On imaging, he was found to have a lesion in segment 2-3 of the left lobe of the liver, which measured at least 3 cm in diameter. He was admitted to the hospital to undergo a laparoscopic left lateral liver resection. This procedure involved laparoscopic ultrasonography of the liver and transection of the left liver lobe with endoscopic linear staplers. The staple height of 3.5, 60 mm long, reinforced with an absorbable polymer membrane was used for liver transection to catch the portal branches. This required multiple firings in the liver parenchyma and additional division of some tissue using the Harmonic scalpel. The larger branch in the middle of segment 2-3 and the left hepatic vein were both transected with the novel staple line reinforcement technique. Bleeding or any bile leakage in this area could not be visualized. No drains were left. The patient's postoperative course was uncomplicated, and he was discharged on postoperative day 3. Pathology results showed a cavernous hemangioma of 4.5 cm in diameter. Staple line reinforcement with the absorbable polymer membrane has the potential to decrease staple line hemorrhage and bile leakage.

Abdominal Pain↗

The phylogeny of proteinase 3/myeloblastin, the autoantigen in Wegener's granulomatosis, and myeloperoxidase as shown by immunohistochemical studies on human leukemic cell lines.

Proteinase 3/myeloblastin (Pr3/MBN) is a serine protease found in primary granules of neutrophilic granulocytes and monocytes in man. This enzyme has been identified as the main autoantigen in Wegener's granulomatosis. Pr3/MBN was earlier identified in the promyelocytic cell line HL-60 and was assumed to be at least partly responsible for controlling proliferation and differentiation within the granulocytic-monocytic system. We have investigated 98 leukemic cell lines representing all lineages of the hemopoietic system. Pr3/MBN was found to appear in mature myeloblastic precursors and in CD34+ immature monoblastic cells and was restricted to the granulocytic and monocytic lineage. Myeloperoxidase (MPO) was found to be expressed earlier in development in the granulocytic lineage, but appeared later than Pr3/MBN in the monocytic lineage. The identification of cell lines which contained Pr3 only or MPO only may make them useful for demonstration of anti-neutrophil cytoplasmic antibodies in primary vasculitides.

Autoantigens↗