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PubMed1978

Performance comparison of the Corning thin-film agarose and the Hyland thick-film agar methods for immunoelectrophoresis.

Many clinical laboratories do immunoelectrophoresis for qualitative assessment of proteins in biological fluids. Commercial kits are available that supply some or all of the necessary components, but the nature of these components varies. Seeking a reliable method for most easily preserving the original immunoelectrophoresis pattern as a permanent record, we compared a thick-film agar method to a thin-film agarose method in immunoelectrophoresis of a total of 70 serum and urine samples. For each sample, either method resulted in the same interpretation. The thin-film agarose method not only yields a stained, permanent record in about the same time that the thick film agar is ready for interpretation but not preservation, but also requires less antiserum, a smaller sample, and may not require purchase of additional hardware.

Blood Proteins↗
PubMed1994

Structural organization of the gene for human CD36 glycoprotein.

The cell-surface glycoprotein CD36 interacts with a large variety of ligands, including collagen types I and IV, thrombospondin, erythrocytes parasitized with Plasmodium falciparum, platelet-agglutinating protein p37, oxidized low density lipoprotein, and long-chain fatty acids. Its expression is restricted to platelets, monocytes, adipocytes, and some endothelial and epithelial cells and is regulated during cell activation, differentiation, and development. CD36 belongs to a novel gene family of structurally related glycoproteins that includes CLA-1 and the lysosomal membrane glycoprotein LIMPII. To advance our knowledge on the genomic organization and the regulation of the cellular expression of the genes of this family, we have investigated the structural organization of the human CD36 gene and of its 5'-proximal flanking region. The CD36 gene is encoded by 15 exons that extend more than 32 kilobases on the human genome. Interestingly, the CD36 mRNA 5'-untranslated region is encoded by three exons. The 3'-untranslated region is contained in two exons, whose expression pattern can originate two mRNA forms. The cytoplasmic and transmembrane regions predicted at both terminal ends of the polypeptide chain are encoded by single exons, while the extracellular domain is encoded by 11 exons. The transcription initiation site of the CD36 gene is located 289 nucleotides upstream from the translational start codon. Sequence analysis of the proximal 5'-flanking region of the gene reveals the existence of a TATA box appropriately located with respect to the transcription initiation site and several potential cis-regulatory elements that might contribute to the transcriptional regulation of the CD36 gene. Delineation of the structural organization of the CD36 gene may help in defining the boundaries of relevant structural and/or functional domains in CD36 and, by extension, in the other members of the family.

Antigens, CD↗
PubMed1995

Reorganization of ascending 5-HT axon projections in animals previously exposed to the recreational drug (+/-)3,4-methylenedioxymethamphetamine (MDMA, "ecstasy").

The recreational drug (+/)3,4-methylenedioxymethamphetamine (MDMA, "ecstasy") is a methamphetamine derivative that selectively destroys central 5-HT axons and axon terminals in animals and, possibly, humans. The fate of 5-HT neurons following MDMA injury is uncertain. In particular, while it is known that central 5-HT axons can undergo regenerative sprouting after MDMA injury, it has not been determined whether they reestablish the original innervation pattern. To address this question, the present studies examined 5-HT innervation patterns in animals lesioned with MDMA 12-18 months previously. Both rodents (rats) and nonhuman primates (squirrel monkeys) were examined, since there is indication that serotonergic recovery after MDMA injury may be species dependent. 5-HT axon projections were studied neurochemically, autoradiographically and immunocytochemically. In both rodents and nonhuman primates previously lesioned with MDMA, substantial serotonergic axonal sprouting was observed. However, in a few rats and in most squirrel monkeys, the reinnervation pattern was highly abnormal: distant targets (e.g., dorsal neocortex) remained denervated, while some proximal targets (e.g., amygdala, hypothalamus) were reinnervated or hyperinnervated. Although the specific determinants of axonal recovery after MDMA injury remain to be identified, it appears that axons which initially sustain more severe damage, are longer, or are more highly arborized have low probability of recovering. The observation that some brain regions remain denervated, while others are reinnervated or hyperinnervated suggests that, under some circumstances, MDMA injury can lead to a lasting reorganization of ascending 5-HT axon projections. Such lasting changes in brain innervation, documented here in MDMA-treated animals, may have implications for humans using MDMA recreationally.

Afferent Pathways↗
PubMed1994

The restricted surgical relevance of morphologic criteria to classify systemic-pulmonary collateral arteries in pulmonary atresia with ventricular septal defect.

Now that systemic-pulmonary collateral arteries are used for unifocalization in patients with pulmonary atresia and ventricular septal defect, the question arises whether morphologic criteria of these collateral arteries could help to provide better results. In an attempt to classify the morphologic features of systemic-pulmonary collateral arteries, we studied 31 heart-lung autopsy specimens with pulmonary atresia and ventricular septal defect. The course of the systemic-pulmonary collateral arteries (origin, branching pattern, and connections with systemic and central pulmonary arteries) was related to their histologic characteristics. The results show that systemic-pulmonary collateral arteries cannot be classified according to their course related to the trachea and the main branches of the bronchi. The histologic features of these collateral arteries vary along their course to the lungs. Nearly all systemic-pulmonary collateral arteries contain a muscular or a musculoelastic segment. One type of collateral artery (complex loop anastomoses) is completely muscular and resembles a bronchial artery. Nutritive branches (bronchial arteries) arise from all histologic types of systemic-pulmonary collateral artery segments. The size and number of intimal proliferations in muscular, elastic, and musculoelastic segments did not differ significantly. In 29 of 31 cases a ductus arteriosus did not coexist with large collateral arteries (two cases unknown). It is concluded that a classification of large systemic-pulmonary collateral arteries based on morphologic features results in a highly variable system, which does not facilitate decisions for the suitability of these arteries for unifocalization procedures. The variability of the systemic-pulmonary collateral arteries corresponds with the recent embryologic finding that during development, collateral artery formation is possible during extended periods.

Adolescent↗
PubMed1993

[Changes in the cellular ultrastructure of the gill epithelium in Tilapia under the action of cadmium on the fish].

The ultrastructure of the Tilapia gill epithelium cells was studied under condition of a prolonged (2 months) treatment of these fishes with cadmium in a concentration of 5 mg/l. A decrease in the quantity of chloride cells in the primary gill epithelium, and of respiratory cells in the secondary epithelium was found. The chloride and mucous cells are, respectively, most sensitive and most resistant to the influence of cadmium. The accumulation of lysosomal structures in chloride and respiratory cells was observed, in addition to a reduced surface relief in these and some damage in mitochondria of the former being noticed. A slow development of reparation processes in the tilapia gill epithelium cells was followed after the cancellation of cadmium effect. No restoration of the original ultrastructural pattern of the gill epithelium cells was observed after a 1.5-month inhabitance of Tilapia in clean water.

Animals↗
PubMed1993

Introduction to the techniques of molecular biology.

The purpose of this chapter is to outline some of the common recombinant DNA methods in use today. These techniques are usually employed to isolate a defined portion of the genome, mostly a gene, from an organism or tissue of interest and, thereafter, to characterize the structure and function of this genetic material. To isolate a gene, genomic DNA is extracted from a selected tissue. For a better handling the relatively large DNA molecules are cut into a mixture of fragments by restriction endonucleases. The fragments are then separated from each other according to their size by gel electrophoresis. A procedure called Southern blotting is used to verify the presence of the desired gene in one of the DNA fragments separated on an agarose gel. The DNA fragments are transferred from the gel to a filter whereby the original fragment pattern is maintained. Then, a single-stranded DNA or RNA probe specific for the gene to be isolated is hybridized to its target fragments fixed to the filter. A radioactive or fluorescent tag is attached to the probe for subsequent identification. In cases where only transcribed sequences are to be isolated cytoplasmic messenger RNA (mRNA) is prepared instead of DNA. Analysis of RNA by a technique similar to Southern blotting is termed Northern blotting. Preservation of DNA sequences is usually achieved by DNA cloning. DNA cloning involves the insertion of a DNA fragment into a DNA vector and the stable incorporation of the recombinant DNA into a suitable host. Propagation of the host facilitates the amplification of the recombinant DNA for subsequent analysis.(ABSTRACT TRUNCATED AT 250 WORDS)

Blotting, Northern↗
PubMed1976

[Clinical studies on kinking of the neck-vessels].

A survey is given of the clinical course of 273 patients hospitalised at one of the departments for internal diseases of a general hospital because of cerebral lesions of different pattern and origin. By means of angiography carried out on 60 patients kinking of the extracranial arteries was proved in 18 cases. Ten patients were submitted to reconstructive vascular surgery: 7 of them are symptom-free. Signs, symptoms and possibilities of surgical or conservative treatment are discussed.

Adult↗
PubMedDate not supplied

Endoscopic ultrasonography in the evaluation of leiomyoma and extramucosal cysts of the esophagus.

BACKGROUND/AIMS: Leiomyoma is the most common type of benign esophageal tumor, whereas extramucosal cysts of the esophagus are congenital anomalies frequently asymptomatic in the adult and in most cases detected incidentally on chest x-ray. It is worthwhile considering these conditions together, because they present similar diagnostic and surgical problems. Conventional imaging tests do not lead to a precise diagnosis. The purpose of this study was to evaluate the use of endoscopic ultrasonography in the diagnosis of, and planning of treatment modalities for, these conditions. METHODOLOGY: Fifteen patients with esophageal leiomyoma and seven patients with extramucosal esophageal cysts were studied with endoscopic ultrasonography using an Olympus GF- EU-M3 instrument with a 7.5-12 MHz echoprobe. In all patients, the results of endoscopic ultrasonography were compared with the histology of the resected specimens. RESULTS: The histology of the resected specimens confirmed the endosonographic diagnosis in all patients. No malignancy was found in any specimen. CONCLUSIONS: Endoscopic ultrasonography is very accurate in visualizing these lesions and differentiating cystic from solid submucosal esophageal masses; in addition, the test can establish the exact location of the mass in relation to the esophageal wall and mediastinum. Therefore, endoscopic ultrasonography has a great impact in confirming the diagnosis of leiomyoma and extramucosal cysts of the esophagus and facilitates therapeutic decision-making because of its capacity to clearly define the size, layer of the origin, and pattern of the mass.

Adult↗
PubMed1995

Characterization of a novel human IgG antibody reactive with a Ca(2+)-sensitive cell-cell adhesion epitope of PtK2 epithelial cells.

We have characterized a human IgG antibody present in the serum of a patient with an autoimmune undifferentiated connective tissue disease and reactive with PtK2 epithelial cell-cell adhesions. The fluorescent staining pattern is observed only at cell-cell contacts whether cells are permeabilized or not. The serum reacts with polypeptides of 90, 48 and 45 kD by immunoblotting. IgG affinity-purified from these bands failed to reproduce the original immunofluorescence staining pattern. Treatment with cycloheximide did not abolish the staining pattern suggesting that the recognized antigen is not a newly expressed protein. However, when EGTA was used for chelating calcium ions in the culture medium the original staining pattern observed at cell-cell adhesions was affected although some fluorescence was still present at cell periphery. This was reversible when cells were reincubated with fresh medium containing Ca2+. The recognized antigen colocalizes at cell-cell adhesions with actin, the microfilament-associated proteins vinculin, alpha-actinin and myosin light chain, and with Triton-insoluble uvomorulin (E-cadherin) material. We conclude that the antibody reacts with, at least, an extracellular portion of a Ca(2+)-dependent PtK2 antigen. The characterization of this antibody based on (1) its localization at cell-cell adhesions, (2) its sensitivity to EGTA-treatment and (3) its colocalization with the epithelial cellular adhesion molecule (CAM) uvomorulin, strongly suggest that the recognized Ag is a CAM or a CAM-associated protein.

Autoimmune Diseases↗
PubMed1998

[The role of the interatrial septum in development of supraventricular tachyarrhythmias of vagal origin in dogs].

Patterns of atrial activation of ectopic pulse source during vagal stimulation were studied on atrial epicardial surface and atrial septum in mongrel dogs. Epicardial activation maps were drafted. The findings show that, irrespective of preferred direction of conduction in the atria and septum, the entire atrial myocardium acted as a conducting system. Mapping of the first extrasystolic beats during tachyarrhythmias induced with vagal stimulation, has shown their ectopic origin. The data obtained confirms the suggestion that atrial fibrillation may be due to a single source of arrhythmia in septal area.

Animals↗
PubMed2001

SOM integrated with CCA for the feature map and classification of complex chemical patterns.

Considering that the two-dimensional (2D) feature map of the high-dimensional chemical patterns can more concisely and efficiently represent the pattern characteristic, a new procedure integrating self-organizing map (SOM) networks with correlative component analysis (CCA) is proposed. Firstly, CCA was used to identify the most important classification characteristics (CCs) from the original high-dimensional chemical pattern information. Then, the SOM maps the first several CCs, which include the most useful information for pattern classification, onto a 2D plane, on which the pattern classification feature is concisely represented. To improve the learning efficiency of SOM networks, two new algorithms for dynamically adjusting the learning rate and the range of neighborhood around the winning unit were further worked out. Besides, a convenient method for detecting the topologic nature of SOM results was proposed. Finally, a typical example of mapping two classes natural spearmint essence was employed to verify the effectiveness of the new approach. The feature-topology-preserving (FTP) map obtained can well represent the classification of original patterns and is much better than what obtained by SOM alone.

Journal Article↗
PubMed2006

Snail1 gene function during early embryo patterning in mice.

Originally identified as one of two zygotically expressed genes required for gastrulation in Drosophila, the Snail gene and other family members play critical roles in vertebrate development. Functionally, these genes are thought to drive epithelial-mesenchymal transitions at several points during development, and also during the metastatic progression of cancer. Although the Snai2-null mouse is viable and fertile, the early embryonic lethality of Snai1-null mice has precluded the detailed analysis of Snai1 function after gastrulation. We have recently generated a conditional allele of the Snai1 gene and examined its function during the formation of the neural crest and establishment of the left-right axis. We uncovered new details regarding Snai1 function during gastrulation and left-right asymmetry determination, while surprisingly showing that neither the Snai1 nor Snai2 genes are essential for neural crest cell delamination. These results shed new light on the role of Snail family genes in early mouse development, and raise interesting questions concerning the diversity of gene function among vertebrate species.

Alleles↗
PubMed2006

Asymmetric expression of the BMP antagonists chordin and gremlin in the sea anemone Nematostella vectensis: implications for the evolution of axial patterning.

The evolutionary origin of the anterior-posterior and the dorsoventral body axes of Bilateria is a long-standing question. It is unclear how the main body axis of Cnidaria, the sister group to the Bilateria, is related to the two body axes of Bilateria. The conserved antagonism between two secreted factors, BMP2/4 (Dpp in Drosophila) and its antagonist Chordin (Short gastrulation in Drosophila) is a crucial component in the establishment of the dorsoventral body axis of Bilateria and could therefore provide important insight into the evolutionary origin of bilaterian axes. Here, we cloned and characterized two BMP ligands, dpp and GDF5-like as well as two secreted antagonists, chordin and gremlin, from the basal cnidarian Nematostella vectensis. Injection experiments in zebrafish show that the ventralizing activity of NvDpp mRNA is counteracted by NvGremlin and NvChordin, suggesting that Gremlin and Chordin proteins can function as endogenous antagonists of NvDpp. Expression analysis during embryonic and larval development of Nematostella reveals asymmetric expression of all four genes along both the oral-aboral body axis and along an axis perpendicular to this one, the directive axis. Unexpectedly, NvDpp and NvChordin show complex and overlapping expression on the same side of the embryo, whereas NvGDF5-like and NvGremlin are both expressed on the opposite side. Yet, the two pairs of ligands and antagonists only partially overlap, suggesting complex gradients of BMP activity along the directive axis but also along the oral-aboral axis. We conclude that a molecular interaction between BMP-like molecules and their secreted antagonists was already employed in the common ancestor of Cnidaria and Bilateria to create axial asymmetries, but that there is no simple relationship between the oral-aboral body axis of Nematostella and one particular body axis of Bilateria.

Amino Acid Sequence↗
PubMed1989

Polarity and patterning in the neural tube: the origin and function of the floor plate.

Little is known about the cellular and molecular mechanisms that determine neuronal cell fate and the patterning of neuronal connections in the vertebrate central nervous system. In this paper we summarize evidence which indicates that some aspects of neuronal differentiation and axon guidance are regulated by specialized epithelial cells that occupy the medial region of the neural plate and, later, the ventral midline of the spinal cord. This cell group, termed the notoplate/floor plate appears to constitute a distinct compartment within the neural plate that is more closely related in lineage and perhaps also in function to axial mesodermal cells of the underlying notochord than to other neural plate cells. Cells of the notoplate exhibit specialized mechanical and adhesive properties that may contribute to neurulation. At later stages of development, the floor plate appears to guide developing axons in the embryonic spinal cord by releasing a diffusible chemoattractant factor and by virtue of its specialized cell surface properties. The floor plate may also play a role in the determination of cell identity and patterning at earlier stages of neural tube development.

Animals↗
PubMed1994

HLA-DQB1*0305 and -DQB1*0304 alleles among Sardinians. Evolutionary and practical implications for oligotyping.

This study, performed in individuals of Sardinian descent, reports an epidemiologic and molecular analysis of the recently identified DQB1*0304 and DQB1*0305 alleles. These two alleles having a gene frequency of 0.017 and 0.005, respectively, are not uncommon in Sardinia and are distributed fairly uniformly on the island. The analysis of DQB1 second and third exons of the two alleles revealed that although they have always been found included within the same DRB1*0403-DQA1*03 haplotype, they had a different origin. The sequence pattern of DQB1*0305 confirmed that it originated from the DQB1*0302 "recipient" gene by the insertion of a DQB1*0402 nucleotide stretch, within its beta-sheet region, while that of DQB1*0304 suggested that it originated from the DQB1*0301 gene, either by a single point mutation at codon 57 (GCC instead of GAC) or, alternatively, by a segmental transfer of a DQB1*0302 motif, including codon 57, within its alpha-helic region. Independently from the mechanism of generation, the fact that DQB1*0304 originated from DQB1*0301 allele was intriguing considering that, in over 1500 HLA class II Sardinian haplotypes examined, neither the putative parental DRB1*0403-DQA1*03-DQB1*0301 haplotypes were found. Finally, since the assignment of DQB1*0305 may be inaccurate with the traditional panel of probes commonly used for DQB1 oligotyping, the use of an additional oligonucleotide probe is recommended.

Alleles↗
PubMed2001

Variability of the lipolytic activity in Yarrowia lipolytica and its dependence on environmental conditions.

This work was aimed to the evaluation of the variability of lipolytic activity in Yarrowia lipolytica strains, as well as to asses for a selected strain, the response to the changes of physico-chemical variables (such as pH, NaCl and lipid content), in order to obtain predictive models describing their effects on the lipolysis pattern. The strains tested, having different environmental origin, showed different patterns of the free fatty acids (FFA) released. The clustering of the free fatty acids profiles evidenced that the unweighted average distance within the strains of the same species did not exceeded 30%. However, the lipolytic activity of some strains generated FFA profiles that differentiated from the majority of the strains considered. Also, when a single strain was inoculated in model systems in which pH, NaCl and milk fat were modulated according to a Central Composite Design (CCD), chemico-physical characteristics of the system led to marked variations in the lipolytic activity with consequent changes in individual fatty acids released. In most cases, when the same Y. lipolytica strain was used, under the experimental conditions adopted, the modulation of the lactic acid, NaCl and lipid content did not generate differences in the fatty acid release exceeding 20-21%. However, some combinations of factors remarkably affected lipase expression or activity, and generated differences in the fatty acid released higher than those observed among different strains of the same species.

Animals↗