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Results for “Cytoplasmic Streaming”

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PubMed1962

An electron microscopic study of the passage of colloidal particles from the blood vessels of the ciliary processes and choroid plexus of the rabbit.

The thinnest areas of the capillaries of the choroid plexus and ciliary processes in the eye of the rabbit are characterized by the presence of fenestrae. When various colloidal particles opaque to the electron beam (thorotrast, gold sol, and saccharated iron oxide) were injected into the blood stream, none were found in fenestrae or in areas that might suggest their having passed through fenestrae. The passage of marker particles from the lumen to the surrounding connective tissue does take place on occasion in the areas of thicker walls in the capillaries and venules rather than in the attenuated and fenestrated endothelial walls. The pathway taken by these markers may be either through the cytoplasm of the endothelial cells via membrane-bounded vesicles and vacuoles or through the intercellular spaces of the vessels. An altered aqueous humor (cloudy and plasmoid) was produced by endotoxin injection or by making a draining fistula in rabbit cornea. Both methods gave rise to the same changes in the blood vessels of the ciliary processes. Under such conditions of inflammation the passage of colloidal particles through the thicker walls of the capillaries and venules was greatly increased and occurred primarily as an intercellular passage between the endothelial cells. The attenuated and fenestrated areas of the endothelium of the small capillaries remained unchanged with no particles passing through them. These results on the altered vessels of the ciliary processes parallel those of Majno and Palade (26) on the rat cremaster muscle.

Animals↗

PubMed1991

Connectional studies of the primate lateral geniculate nucleus: distribution of axons arising from the thalamic reticular nucleus of Galago crassicaudatus.

Anterograde and retrograde transport methods have been used to explore the interconnections between the thalamic reticular nucleus (TRN) and the dorsal lateral geniculate nucleus of Galago crassicaudatus. We first defined the region of the TRN, which is connected to the lateral geniculate nucleus, by examining the distribution of geniculo-TRN axons, cortico-TRN axons arising from area 17, and the location of TRN-geniculate neurons. Following an intraocular injection of 3H-proline/3 H-leucine, trans-synaptically transported protein is present bilaterally within the lateral portion of the caudal TRN. This same caudal and lateral region is also targeted by cortico-TRN axons and contains neurons which project upon the lateral geniculate nucleus. Light microscopic anterograde transport methods were used to analyze the distribution of TRN-geniculate axons. Our data reveal that all layers and interlaminar zones of the dorsal lateral geniculate nucleus contain TRN axons. Electron microscopic-autoradiographic data support and extend our light microscopic findings by revealing labeled TRN terminals within all geniculate layers. These TRN profiles are the same size throughout the geniculate and exhibit morphological characteristics similar to F1 terminals described by others. That is, they possess predominantly pleomorphic vesicles, a dark cytoplasmic matrix, dark mitochondria, and symmetrical synaptic contacts. Two additional features of TRN terminals have been observed in some profiles. These include dense-core vesicles and a dense, punctate cytoplasmic matrix, which is sometimes associated with the postsynaptic specialization. In addition to their morphology and size, the postsynaptic targets of TRN terminals are similar within the three sets (parvi-, magno-, and koniocellular) of geniculate layers. TRN profiles terminate upon dendrites of all sizes and somata. These findings suggest that the TRN modulates the retino-geniculocortical pathway and that this modulation is occurring in all three streams.

Animals↗

PubMed2006

Classic cadherins regulate tangential migration of precerebellar neurons in the caudal hindbrain.

Classic cadherins are calcium dependent homophilic cell adhesion molecules that play a key role in developmental processes such as morphogenesis, compartmentalization and maintenance of a tissue. They also play important roles in development and function of the nervous system. Although classic cadherins have been shown to be involved in the migration of non-neuronal cells, little is known about their role in neuronal migration. Here, we show that classic cadherins are essential for the migration of precerebellar neurons. In situ hybridization analysis shows that at least four classic cadherins, cadherin 6 (Cad6), cadherin 8 (Cad8), cadherin11 (Cad11) and N-cadherin (Ncad), are expressed in the migratory streams of lateral reticular nucleus and external cuneate nucleus (LRN/ECN) neurons. Functional analysis performed by electroporation of cadherin constructs into the hindbrain indicates requirement for cadherins in the migration of LRN/ECN neurons both in vitro and in vivo. While overexpression of full-length classic cadherins, NCAD and CAD11, has no effect on LRN/ECN neuron migration, overexpression of two dominant negative (DN) constructs, membrane-bound form and cytoplasmic form, slows it down. Introduction of a DN construct does not alter some characteristics of LRN/ECN cells as indicated by a molecular marker, TAG1, and their responsiveness to chemotropic activity of the floor plate (FP). These results suggest that classic cadherins contribute to contact-dependent mechanisms of precerebellar neuron migration probably via their adhesive property.

Animals↗

PubMed2005

Effect of caspase 9 related signaling molecules on the apoptosis of human vascular endothelial cell induced by homocysteine.

OBJECTIVE: To understand the role of mitochondria associated signaling pathway in the apoptosis of human vascular endothelial cell induced by homocysteine (Hcy). METHODS: The mRNA and protein expression levels of the up-stream signaling molecules of caspase 3, Bcl 2, caspase 9, and cytosolic cytochrome-c, were investigated. The in vitro cultured human umbilical vein endothelial cells with homocysteine at different concentrations were incubated for 24 h. The expressions of Bcl 2 and caspase 9 at mRNA and protein levels were analyzed by reverse transcription-polymerase chain reaction(RT-PCR) and Western blot. Cytochrome-c in cytoplasm was also detected by Western blot. RESULTS: The expression levels of three signaling molecules were all down-regulated by homocysteine at both mRNA and protein levels in a dose-dependent manner. CONCLUSION: Homocysteine could affect the formation of apoptosome through repressing the expression of Bcl 2 gene and release of cytochrome-c from mitochondria. Decreasing of apoptosome could disturb the activation of caspase 9. The results also indicate that the mitochondria pathway is not the major signaling pathway involved in Hcy-induced apoptosis.

Apoptosis↗

PubMed1978

Brain macrophages in rats following intravenous labelling of mononuclear leucocytes with colloidal carbon.

Intravenous injections of colloidal carbon were used to label circulating mononuclear leucocytes. In the neonatal rats (3-5 days old), either 24 or 48 hours later, carbon-labelled macrophages were seen in the brain tissue. In the areas examined, notably the corpus callosum and the cerebral cortex, labelled macrophages were distributed randomly. They were either perivascular, perineuronal or lay between the nerve fibres. The labelled cells were mostly spindle-shaped with an eccentric nucleus and the cytoplasm at one pole of the cell was engorged with dark carbon particles. Abundant labelled cells were also seen over the brain surface in the layers of meninges. There was no evidence of leucocytic infiltration into the brain tissue of mature animals. It is concluded from the present work that a proportion (but not all) of the macrophages in the neonatal rat brain are derived from the blood stream.

Age Factors↗

PubMed1983

Vascular changes in nerves in experimental leprosy--an ultrastructural study.

Ultrastructural observations made on the blood vessels of nerves in CBA/J mice with experimentally produced M. leprae infection revealed cytoplasmic filamentous structures along with large sized vacuoles in the endothelial cells and reduplication of peripericytal basement membranes in the 6-9 months of infection. In animals with late infection of 9-15 months duration, the endothelial cells presented foamy appearance caused by an increase in the number of vacuoles. These findings are suggestive of degenerative changes in the endothelial cells probably due to circulation of noxious substance in the blood stream.

Animals↗

PubMed1978

Scanning electron microscopy (s.e.m.) of biopsy specimens of ruptured intracranial saccular aneurysms.

Surgical specimens of 4 intracranial saccular aneurysms were studied by scanning electron microscopy. The internal surface of the aneurysms showed crater-like defects and cytoplasmic bridges. In some areas the endothelium was preserved, but its longitudinal convolutions were higher and thicker than those found in unchanged areas. On the damaged endothelial surface there was an increased number of blood cells. The adventitia resembled that of a normal cerebral artery. In conclusion the alterations observed are similar to those found in atherosclerosis and are most likely due to the high wear and tear provoked by the blood streaming into the aneurysm. The results of this study are in agreement with the findings of a companion transmission electron microscopy study and emphasize the importance of degenerative changes on the development and rupture of intracranial saccular aneurysms.

Adult↗

PubMed2005

Nuclear phospholipase C: involvement in signal transduction.

During the past years, several independent laboratories have highlighted the presence of nuclear signaling pathways based on lipid hydrolysis, which are not a mere duplication of those occurring at the plasma membrane. Among the enzymes of the cycle, nuclear phosphoinositide-specific phospholipase C (PI-PLC) has been analyzed quite extensively. In this context, PI-PLCbeta1 appears to play a key role as a check point in the G1 phase of the cell cycle. It has also been shown that its activation and/or up-regulation is upon the control of type 1 insulin-like growth factor receptor (IGF-R) in both mouse fibroblast and myoblasts, suggesting that its signaling activity is essential for the normal behavior of the cell, at least in culture. The recent discovery of a possible involvement of the deletion of PI-PLCbeta1 gene in the progression of myelodysplastic syndrome (MDS) to acute myeloid leukemia (AML) in humans strengthens the contention that nuclear PI-PLC signaling is essential for physiological processes such as cell growth and differentiation. Even though PI-PLCbeta1 is present and does not translocate to eukaryotic nuclei, this organelle, even though only in some conditions contains also PI-PLCgamma1 which acts not only as a PI-PLC but also as guanine nucleotide exchange factor (GEF) for PI 3-kinase enhancer (PIKE) and is somehow linked to PI 3-kinase (PI3K) activity. Also members of PI-PLCdelta family are shuttling from the nucleus to the cytoplasm and return and are possibly involved in the control of cell growth. We must also take into account the presence in the nucleus of other phospholipases such as phospholipase A2 (PLA2) and phospholipase D (PLD), which also exert a signaling activity upon external stimuli. On the whole this review highlights the latest development in the PI-PLC cycle in the nucleus, which in terms of activation, regulation and down-stream targets differs substantially from that located at the plasma membrane.

Active Transport, Cell Nucleus↗

PubMed2003

Human mitochondrial C1-tetrahydrofolate synthase: gene structure, tissue distribution of the mRNA, and immunolocalization in Chinese hamster ovary calls.

C1-tetrahydrofolate (THF) synthase is a trifunctional enzyme found in eukaryotes that contains the activities 10-formyl-THF synthetase, 5,10-methenyl-THF cyclohydrolase, and 5,10-methylene-THF dehydrogenase. The cytoplasmic isozyme of C1-THF synthase is well characterized in a number of mammals, including humans; but a mitochondrial isozyme has been previously identified only in the yeast Saccharomyces. Here, we report the identification and characterization of the human gene encoding a functional mitochondrial C1-THF synthase. The gene spans 236 kilobase pairs on chromosome 6 and consists of 28 exons plus one alternative exon. The gene encodes a protein of 978 amino acids, including an N-terminal mitochondrial targeting sequence. The mitochondrial isozyme is 61% identical to the human cytoplasmic isozyme. Expression of the gene was detected in most human tissues, but transcripts were highest in placenta, thymus, and brain. Two mRNAs were detected, a 3.6-kb transcript and a 1.1-kb transcript, and both transcripts were observed in varying ratios in each tissue. The shorter transcript results from an alternative splicing event, where exon 7 is spliced to exon 8a instead of exon 8. Exon 8a is derived from an exonized Alu sequence, sharing no homology with exon 8 of the long transcript, and encodes just 15 amino acids followed by a stop codon and a polyadenylation signal. This short transcript potentially encodes a bifunctional enzyme lacking 10-formyl-THF synthetase activity. Both transcripts initiate at the same 5'-site, 107 nucleotides up-stream of the ATG start codon. The full-length (2934 bp) cDNA fused to a C-terminal V5 epitope tag was expressed in Chinese hamster ovary cells. Immunoblots of subfractionated cells revealed a 107-kDa protein only in the mitochondrial fractions of these cells, confirming the mitochondrial localization of the protein. Yeast cells expressing the full-length human cDNA exhibited elevated 10-formyl-THF synthetase activity, confirming its identification as the human mitochondrial C1-THF synthase.

Alternative Splicing↗

PubMed2002

Transplantation of human neural progenitor cells into the neonatal rat brain: extensive migration and differentiation with long-distance axonal projections.

Here we examined the ability of human neural progenitors from the embryonic forebrain, expanded for up to a year in culture in the presence of growth factors, to respond to environmental signals provided by the developing rat brain. After survival times of up to more than a year after transplantation into the striatum, the hippocampus, and the subventricular zone, the cells were analyzed using human-specific antisera and the reporter gene green fluorescent protein (GFP). From grafts implanted in the striatum, the cells migrated extensively, especially within white matter structures. Neuronal differentiation was most pronounced at the striatal graft core, with axonal projections extending caudally along the internal capsule into mesencephalon. In the hippocampus, cells migrated throughout the entire hippocampal formation and into adjacent white matter tracts, with differentiation into neurons both in the dentate gyrus and in the CA1-3 regions. Directed migration along the rostral migratory stream to the olfactory bulb and differentiation into granule cells were observed after implantation into the subventricular zone. Glial differentiation occurred at all three graft sites, predominantly at the injection sites, but also among the migrating cells. A lentiviral vector was used to transduce the cells with the GFP gene prior to grafting. The reporter gene was expressed for at least 15 weeks and the distribution of the gene product throughout the entire cytoplasmic compartment of the expressing cells allowed for a detailed morphological analysis of a portion of the grafted cells. The extensive integration and differentiation of in vitro-expanded human neural progenitor cells indicate that multipotent progenitors are capable of responding in a regionally specific manner to cues present in the developing rat brain.

Animals↗

PubMed2003

Immunocytochemical localization of reelin in the olfactory bulb of the heterozygous reeler mouse: an animal model for schizophrenia.

Because heterozygous reeler (HR) mice share some abnormal traits with schizophrenic patients, and schizophrenia is often accompanied by impairment of olfactory function, this study examines reelin in the olfactory bulb of the HR mouse. In the WT mouse, reelin immunoreactivity is found in the extracellular matrix, and in the cytoplasm of olfactory nerve fibers, GABAergic interneurons, and glutamatergic mitral cells. Western blot analysis reveals that reelin immunoreactivity in the HR mouse is reduced by 45% compared to WT mouse. This is especially evident in the glomerular GABAergic interneurons. In WT mitral cells, reelin is found in discrete clumps near the axon hillock and within the axon. In the HR mouse, reelin axonal staining is diffuse and densely packed. In the rostral migratory stream of the HR mouse, immunolabeling shows an accumulation of reelin-containing neuronal precursors, apparently unable to shift from tangential to radial migration. These observations indicate that there is a downregulation of reelin in the HR mouse and suggest that secretion of reelin may be compromised. Further studies of the HR mouse may provide a new basis for understanding the role of reelin in the adult CNS, especially as it may relate to schizophrenia.

Animals↗

PubMed1998

Glucagon-like peptide-1 (GLP-1): a gut hormone of potential interest in the treatment of diabetes.

GLP-1 (glucagon-like peptide-1) is a gut hormone which is released into the blood stream after feeding. Its main action is to stimulate insulin secretion through potentiating the insulinotropic action of glucose. The peptide is encoded in the glucagon gene and expressed mainly in the gut L cells. It exerts its actions through activating specific receptors of the seven transmembraneous domain-G-protein-coupled type with 463 amino acids. Its main signalling mechanism is activation of adenylate cyclase and formation of cyclic AMP. The peptide also increases the cytoplasmic concentration of Ca2 which is thought to be executed both through a Na(+)-dependent uptake of extracellular Ca2+ and through release of Ca2+ from intracellular Ca2+ stores. GLP-1 also inhibits glucagon secretion and inhibits gastric emptying and gastric acid and pancreatic exocrine secretion. Its integrated action on carbohydrate metabolism results in reduction of circulating glucose, and GLP-1 has therefore been suggested as a therapeutic alternative in diabetes. Finally, GLP-1 is also expressed in neurons in the hypothalamus, and may be involved in the regulation of feeding behaviour, since it inhibits food intake.

Adenylyl Cyclases↗

PubMed1985

Ultrastructural observations on sinusoidal endothelial cells in chronic active hepatitis.

Ultrastructural features of 12 liver biopsies from patients with chronic active hepatitis were studied, particular attention being paid to endothelial cells. In areas of piecemeal necrosis and parenchymal inflammation sinusoidal endothelial cells show swelling of the cytoplasm, protrusion of the cell body into the sinusoidal lumen, increase in micropinocytotic vesicles and appearance of numerous dense bodies. This cell type is termed 'active endothelial cell'. Subsequent changes include enlargement of the Golgi complex, increase of rough endoplasmic reticulum in cytoplasmic processes with concomitant decrease of dense bodies, appearance of a fuzzy coat and formation of hemidesmosomes in close relationship to basement membrane-like material and reticulin fibres in the space of Disse. The latter ultrastructural characteristics correspond to those of 'fibroblastic reticulum cells' described in lymph nodes. Active endothelial cells and fibroblastic reticulum cells may play a protective role in liver parenchymal inflammation by reducing the accessibility of noxious agents from the blood stream to liver parenchymal cells, and be crucial in the initiation of perisinusoidal fibrosis.

Biopsy, Needle↗

PubMed1981

Morphology and distribution of cytoplasmic inclusions in adrenoleukodystrophy.

We report some additional observations on the pathognomonic cytoplasmic inclusions in a typical case of adrenoleukodystrophy (ALD). Study of the linear inclusions by means of tilt-stage electron microscopy indicated that they represent trilaminar sheet-like structures distinct from materials seen in various other storage disorders. This study emphasizes that the inclusions in the adrenal and testicular parenchymal cells morphologically differ from those in the brain macrophages. In the latter they occur as membrane-bound discrete structures while in the parenchymal cells they are always seen lying free in the cytoplasm and associated with cleft-like spaces. This morphologic variation of the inclusions is noteworthy and probably reflects a biochemical difference in the storage materials in these two most severely affected organs. In addition, this study demonstrates the presence of linear inclusions within macrophages in otherwise normal-appearing liver, spleen, thymus and also in the renal tubular cells. These findings suggest that the abnormal materials containing very long chain fatty acids enter the blood stream and become sequestered in the reticuloendothelial system and possibly are also excreted in the urine. Detection of these abnormal fatty acids in blood and urine may simplify diagnosis of ALD.

Adolescent↗

PubMed1996

Transcriptional and post-transcriptional regulation of pr22 (Op18) with proliferation control.

The pr22 gene was isolated as a gene which is expressed in proliferating cells but not in cells which are differentiated or growth-arrested. When cells of the human monocytic cell line, U937, were differentiated into macrophages, transcription of pr22 was almost completely suppressed. Serum starvation resulted in the inhibition of transcription, although U937 failed to differentiate. In a culture synchronized with excess thymidine, mRNA of pr22 was detected at the G1/S boundary, with the level increasing in the S phase and decreasing in the G2 phase. The gene product, pr22 protein (Pr22) was found to be identical to Op18 as well as to a catastrophe factor. Genes homologous to pr22 were detected in the genome of mouse but not in that of yeast, or Drosophila. The 5' up-stream region of the genomic pr22 contained CpG islands but no TATA box at its appropriate position. About 20% of cell nuclei of normal human fibroblasts were stained in a speckled manner with a monoclonal antibody for C-terminal peptide of Pr22, and these cells were found to be in phases S and G2. The mitotic apparatus was also strongly stained. By Western blot analysis, Pr22 was detected in the nuclear fraction but not in the cytoplasm. The level increased from middle S to G2 phase and remained high until the early G1 phase. N-terminal truncated Pr22 was also detected in these phases. These results suggest that Pr22 may have an additional role other than just functioning in association with microtubules.

Animals↗

PubMed2003

A modified Escherichia coli protein production strain expressing staphylococcal nuclease, capable of auto-hydrolysing host nucleic acid.

The large-scale production of recombinant biotherapeutics, particularly recombinant proteins, provides significant process and regulatory challenges to the biotechnology industry in order to meet the regulatory agencies stringent requirements in a cost-effective manner. Host cell derived nucleic acid causes problems from both a process and a regulatory perspective, as high molecular weight chromosomal DNA is responsible both for the viscosity of cell lysates, and it is a source of heterologous DNA sequences whose inclusion in the final product must be prevented. We have constructed a modified Escherichia coli JM107 expression host (JMN), containing a staphylococcal nuclease expression cassette, integrated into the host chromosome at the dif locus. The nuclease is expressed as a fusion to the ompA signal peptide, and is translocated to the periplasm of the cell, protecting the cytoplasmic nucleic acid from any toxic activity. The nuclease is released during cell lysis, where it subsequently acts to hydrolyse host nucleic acid present in the lysate. Results with this strain show that sufficient levels of nuclease activity are produced to completely auto-hydrolyse the host's chromosomal DNA to a size non-visible on 1% agarose gel, generating a markedly lower lysate viscosity. This provides a suitable methodology to remove heterologous DNA sequences early in the product stream and decrease lysate viscosity, improving the efficiency of downstream processing and product yield, whilst avoiding the addition of exogenous nuclease and its prohibitive costs at large-scale.

Base Sequence↗

PubMedDate not supplied

Transport of microperoxidase across segments of cerebral arterioles under normal conditions.

Microperoxidase, injected intravenously into mice, was allowed to circulate in the blood stream for 1 min to 1 h. The amount of reaction product in the walls of cerebral arterioles was most pronounced after circulation of 40 mg peroxidase for one min, the maximum dose used. The peroxidase was transported across the endothelium in a few segments of cerebral arterioles, lying on the surface of the brain or restricted to regions of the brain parenchyma. Reaction product was observed in vesicles: 1 open to the vessel lumen, 2 freely situated in the cytoplasm of endothelial cells, and 3 open to the subendothelial basement membrane. The endothelial cells did not show leaky plasma membranes. Channels or chains or vesicles, simultaneously open to the vessel lumen and the subendothelial basement membrane, were not seen, nor were fenestrae found. Furthermore, reaction product never formed a continuous column from the first luminal to the first abluminal tight junctions between two adjacent endothelial cells. From these observations, it is suggested that vesicular transfer of intravenously injected microperoxidase may occur under normal conditions across the endothelium in segments of cerebral arterioles.

Animals↗

PubMed1994

Plasmids for heavy metal resistance in Alcaligenes eutrophus CH34: mechanisms and applications.

Alcaligenes eutrophus CH34 is the main representative of a group of strongly related strains (mostly facultative chemolithotrophs) that are well adapted to environments containing high levels of heavy metals. It harbors the megaplasmids pMOL28 and pMOL30 which carry resistance determinants to Co2+, Ni2+, CrO(4)2-, Hg2+, Tl+, Cd2+, Cu2+ and Zn2+. Among the best characterized determinants are the cnr operon (resistance to Co, Ni) on pMOL28 and the czc operon on pMOL30 (resistance to Co, Cd and Zn). Although the two systems reveal a significant degree of amino acid similarity in the structural genes, the regulation of the operons is different. The resistance mechanism in both cases is based on efflux. The efflux mechanism leads to a pH increase outside of the cytoplasmic membrane. Metals are sequestered from the external medium through the bioprecipitation of metal carbonates formed in the saturated zone around the cell. This latter phenomenon can be exploited in bioreactors designed to remove metals from effluents. The bacteria are immobilized on composite membranes in a continuous tubular membrane reactor (CTMR). The effluent continuously circulates through the intertubular space, while the external surface of the tubes is in contact with the growth medium. Metal crystals are eventually removed by the effluent stream and collected on a glass bead column. The system has been applied to effluents containing Cd, Zn, Co, Ni and Cu. By introducing catabolic plasmids involved in the aerobic degradation of PCBs and 2,4-D into metal-resistant A. eutrophus strains, the application range was widened to include effluents polluted with both organic and inorganic substances. Biosensors have been developed which are based on the fusion of genes induced by metals to a reporter system, the lux operon of Vibrio fischeri. Bacterial luciferases produce light through the oxidation of fatty aldehydes. The gene fusions are useful both for the study of regulatory genes and for the determination of heavy metal concentrations in the environment.

Alcaligenes↗