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PubMed2026

Tazarotene-Induced Gene 2 Promotes Melanoma Cell Death via the Activation of Endoplasmic Reticulum Stress.

BACKGROUND: Tazarotene-induced gene 2 (TIG2), also known as retinoic acid receptor responder 2 (RARRES2), encodes the secreted protein TIG2, also known as chemerin, which is involved in immune regulation and metabolism. However, its role in melanoma remains unclear. METHODS: TIG2 expression was analyzed using The Cancer Genome Atlas, Genotype-Tissue Expression, OncoDB, and melanoma tissue cDNA arrays. To evaluate its effects on cell viability and death, TIG2 was overexpressed in A2058 and A375 melanoma cells. RNA sequencing (RNA-seq), qPCR, and Western blotting were performed to identify TIG2-regulated genes and signaling pathways. The involvement of chemokines and endoplasmic reticulum (ER) stress was further examined using the C-X-C motif chemokine ligand 10 (CXCL 10)/CXCL11 and the ER stress inhibitor tauroursodeoxycholic acid (TUDCA). RESULTS: TIG2 expression was reduced in melanoma and other skin cancers. TIG2 overexpression significantly reduced cell viability and induced cell death. RNA-seq analysis showed that TIG2 downregulated CXCL10, CXCL11, and CCL2 while upregulating ER stress-related genes such as HERPUD1 and DDIT3. Exogenous CXCL10 or CXCL11 did not reverse TIG2-mediated effects, whereas TUDCA partially restored cell viability and reduced cell death. CONCLUSIONS: These findings suggest that TIG2 suppresses melanoma cell growth by activating ER stress and modulating immune-related chemokines, highlighting its potential therapeutic relevance.

Endoplasmic Reticulum Stress↗

PubMed1990

Electronmicroscopic demonstration of HPV in oral warts.

Human Papilloma Virus (HPV) has been demonstrated in a series of benign proliferative lesions of the skin and the mucosae. The virus has also been found in verrucous laryngeal carcinoma and carcinomas of the oral cavity and other organs. DNA hybridization techniques have classified, the HPV into 51 types, some of which seem to be associated with specific lesions. In order to study the intracellular distribution of HPV, we performed ultrastructural analysis with the electron microscope on 14 specimens taken from 7 patients by large excisional biopsy, which had been histologically classified as "fibropapilloma". From each patient specimens were taken from both the clinically evident lesion and the clinically normal surrounding mucosa. The specimens were fixed with glutaraldehyde, washed with cacodylate buffer, post-fixed with potassium ferrocyanide reduced-osmium tetroxide, block stained with uranyl acetate and embedded in EPON 812. The tissues underwent to amylase digestion before the electron microscopic examination. We found a large number of viral particles in both nuclei and cytoplasm, without forming crystal array structures as described typically for the virus of the verruca vulgaris (HPV-2). No significant differences were found between the cells derived from the clinical lesion and those derived from the surrounding mucosa. The passage of viral particles from infected to not yet infected cells through the intercellular space was observed. Of particular interest, we found a high intracytoplasmatic presence of the virus and its clear abundance in the cells surrounding the clinical lesion.

Adult↗

PubMed1990

Immuno- and affinity probes for electron microscopy: a review of labeling and preparation techniques.

Immuno- and affinity probes are widely used in biology and medicine, and are becoming essential tools for the elucidation of cell structure and function. This article reviews and discusses the bewildering array of probes and preparation techniques now available for the investigation of sectioned material by transmission electron microscopy, with critical analysis of their merits. Emphasis is placed on immunogold probes and methods useful for routine preparation, gathering together information that may be used to improve labeling techniques. New data on inert dehydration for the localization of sensitive epitopes without chemical or cryofixation is presented.

Affinity Labels↗

PubMed1983

Pre- and post-synaptic structures in insect CNS: intramembranous features and sites of alpha-bungarotoxin binding.

The central neuropile of thoracic ganglia in the central nervous system (CNS) of the cockroach Periplaneta americana contains synapses with characteristic pre- and post-synaptic membrane specializations and associated structures. These include dense pre-synaptic T-bars surrounded by synaptic vesicles, together with post-synaptic densities of varying electron opacity. Exocytotic release of synaptic vesicles is observed only rarely near presynaptic densities, but coated pits are seen at variable distances from them, and may be involved in membrane retrieval. After freeze-fracture, paralinear arrays of intramembranous articles (IMPs) are detected on the P face of many presynaptic terminals, with associated dimples indicative of vesicular release. The E face of these membranes exhibits protuberances complementary to the P face dimples, as well as scattered larger IMPs. Post-synaptic membranes possess dense IMP aggregates on the P face, some of which may represent receptor molecules. Electrophysiological studies with biotinylated alpha-bungarotoxin reveal that biotinylation does not inhibit the pharmacological effectiveness of the toxin in blocking acetylcholine receptors on an identified motoneurone in the metathoracic ganglion. Preliminary thin section ultrastructural analysis of this tissue post-treated with avidin-HRP or avidin-ferritin indicates that alpha-bungarotoxin-binding sites are localized at certain synapses in these insect thoracic ganglia.

Animals↗

PubMed1990

slit: an extracellular protein necessary for development of midline glia and commissural axon pathways contains both EGF and LRR domains.

The Drosophila slit locus encodes a protein with four regions containing tandem arrays of a 24-amino-acid leucine-rich repeat (LRR) with conserved flanking sequences (flank-LRR-flank surrounding these arrays), followed by two regions with epidermal growth factor (EGF)-like repeats. Each of these motifs has been implicated in protein-protein interactions as part of an extracellular domain in a variety of other proteins. Analysis of slit cDNA clones reveals that as a consequence of alternative splicing, the locus can code for two distinct protein species differing by 11 amino acids at the carboxyl terminus of the last EGF repeat. The existence of a putative signal sequence and the absence of a transmembrane domain suggest that slit is secreted, an observation supported by an analysis of its expression in tissue culture. Examining the expression pattern of slit in the embryo by antibody staining, enhancer trap detection, and in situ hybridization, we demonstrate that the protein is expressed by a subset of glial cells along the midline of the developing central nervous system. Through immunoelectron microscopy, slit can be seen on the commissural axons traversing the glial cells although it is absent from the cell bodies of these neurons, implying that slit is exported by the glia and distributed along the axons. Finally, we demonstrate that a reduction in slit expression results in a disruption of the developing midline cells and the commissural axon pathways. The embryonic localization, mutant phenotype, and homology of slit to both receptor-binding EGF-like ligands and adhesive glycoproteins suggest that it may be involved in interactions between the midline glial cells, their extracellular environment, and the commissural axons that cross the midline.

Alleles↗

PubMed1991

Design and evaluation of closed-loop feedback control of minimum temperatures in human intracranial tumours treated with interstitial hyperthermia.

The dynamic nature of blood flow during hyperthermia therapy has made the control of minimum tumour temperature a difficult task. The paper presents initial studies of a novel approach to closed-loop control of local minimum tissue temperatures utilising a newly developed estimation algorithm for use with conductive interstitial heating systems. The local minimum tumour temperature is explicitly estimated from the power required to maintain each member of an array of electrically heated catheters at a known temperature, in conjunction with a new bioheat equation-based algorithm to predict the 'droop' or fractional decline in tissue temperature between heated catheters. A closed loop controller utilises the estimated minimum temperature near each catheter as a feedback parameter, which reflects variations in local blood flow. In response the controller alters delivered power to each catheter to compensate for changes in blood flow. The validity and stability of this estimation/control scheme were tested in computer simulations and in closed-loop control of nine patient treatments. The average estimation error from patient data analysis of 21 sites at which temperature was independently measured (three per patient) was 0.0 degree C, with a standard deviation of 0.8 degree C. These results suggest that estimation of local minimum temperature and feedback control of power delivery can be employed effectively during conductive interstitial heat therapy of intracranial tumours in man.

Adult↗

PubMedDate not supplied

Insonation of fixed porcine kidney by a prototype sector-vortex-phased array applicator.

The sector-vortex applicator, an ultrasound phased array with a geometric focus having multiple sectors and tracks, can directly synthesize, without scanning, diffuse focal patterns useful for hyperthermia. A perfused tissue phantom, consisting of an alcohol-fixed porcine kidney with thermocouples placed in the cortex, is insonated by a prototype sector-vortex applicator with 16 sectors and two tracks at an ultrasound frequency of 750 kHz. Steady-state temperature distributions are measured for a wide range of perfusion rates. Results demonstrate that the radius of the heated region can be controlled effectively by choosing the focal mode of the applicator as it is predicted by theoretical analysis.

Animals↗

PubMedDate not supplied

[Ultrastructural demonstration of the human papilloma virus (HPV) in oral proliferative lesions].

The Human Papilloma Virus (HPV) has been found in a series of benign proliferative lesions of the skin and mucosa. The virus has also been found in verrucous laryngeal carcinoma and carcinomas of the oral cavity and other organs. DNA hybridization techniques have made it possible to classify 51 types of HPVs, some appearing to be associated with specific lesions. In order to study the intracellular distribution of HPVs, an ultrastructural morphological analysis was performed with an electron microscope on 10 specimens taken from 5 patients. The specimens were obtained through large excisional biopsy, histologically classified as "fibropapilloma". For each patient specimens were taken from the clinically evident lesion and from the surrounding clinically normal mucosa. The specimens were fixed in glyceraldehyde, washed in a cacodylate buffer, post-fixed in potassium ferrocyanide reduced-osmium tetroxide, stained with uranyl acetate and included in EPON 812. The tissue was subjected to amylase digestion prior to electron microscope examination. A great number of viral particles were found in both the nucleus and the cytoplasm, without forming crystal array structures as typically described for the verruca vulgaris virus (HPV-2). No significant differences could be found between the cells derived from the clinical lesion and those derived from the surrounding mucosa. The passage of viral particles from infected to as yet uninfected cells through the intercellular space could be seen. The high intracytoplasmatic presence of the virus, and its clear abundance in those cells surrounding the clinical lesion, is felt to be of special interest.

Adult↗

PubMed1990

Glial process elongation and branching in the developing murine neocortex: a qualitative and quantitative immunohistochemical analysis.

Cells of astroglial lineage in the murine cerebrum undergo a succession of transformations during prenatal and early postnatal development. The bipolar radial cell, the earliest astroglial form to appear, provides a radially aligned, parallel array of fibers that serves as a guide to neuronal migration. The multipolar astrocyte is the representative of this lineage that persists in the adult cerebrum. The processes of the multipolar astrocytes form a complex reticulum, which is considered critical to the development, function, and maintenance of neural circuits. A monopolar radial cell appears to be transitional between the two. The shift from the radial glial fiber system to a diffuse glial network is achieved largely in the E17-P2 interval in the mouse. This phenomenon has been studied qualitatively and quantitatively by staining cerebral tissue with monoclonal antibody RC2, a specific and sensitive ligand for cells of astroglial lineage in the mouse. Elongation and branching of glial processes contribute to the glial transformation. Elongation of radial fibers occurs under the guidance of other radial glial fibers (fasciculated elongation) or independently of other fibers (nonfasciculated elongation). Fasciculated elongation results in an increase in the density of radial glial fibers that span the cortical layers. Nonfasciculated elongation appears to be associated with process branching. This is the initial event in transformation of the bipolar radial cells to monopolar radial or multipolar cells. Only nonfasciculated elongation is characteristic of processes of the monopolar radial cells and multipolar astrocytes. Branching of the processes of all three cell forms appears to occur both by bifurcation at the elongating tip and by sprouting from the fiber shaft. Elongating fibers are tipped by growth cones that are relatively simple in shape as compared to those observed at the tips of elongating axons. Growth cones at the tips of nonfasciculated fibers are more complex in form than those at the tips of radial fibers elongating in contact with other radial fibers.

Animals↗

PubMed1992

Nucleotide sequence analysis of rheumatoid factors and polyreactive antibodies derived from patients with rheumatoid arthritis reveals diverse use of VH and VL gene segments and extensive variability in CDR-3.

The heavy and light chain nucleotide sequences of 17 monoreactive and polyreactive rheumatoid factors largely derived from the inflamed synovial tissue of two patients with rheumatoid arthritis are described. Some of these sequences have been the subject of a previous report from our laboratories. Additionally, a few rheumatoid factors from the peripheral blood of patients with systemic lupus erythematosus and Sjogren's syndrome as well as a normal individual are included. A review of our previous results as well as the new data provided within this paper lead to the following major conclusions: (1) Rheumatoid factors and polyreactive antibodies derive from a diverse array of VH and VL gene segments; (2) While many rheumatoid factors and polyreactive antibodies are direct or nearly direct copies of germline genes, some show clear evidence of somatic mutation; (3) The CDR3 of all of these antibodies is extraordinarily diverse in length and composition. Certain 'restrictions' do appear in this very large sample: (a) the polyreactive antibodies are exclusively lambda, and (b) there seems to be a preponderance of a particular subset of VH3 genes beyond that one would expect based on random utilization.

Amino Acid Sequence↗

PubMed1986

Short repeats cause heterogeneity at genomic terminus of bovine herpesvirus 1.

Analysis of the genomes of different bovine herpesvirus 1 strains revealed a UL terminal HindIII fragment differing in size (from 2.4 to 2.8 kilobases). This fragment polymorphism occurred in the DNA of a wild-type isolate, in highly passaged, apathogenic tissue culture derivatives, and in plaque-purified substrains. This heterogeneity was due to variations in the copy number of a 14-base-pair tandem repeat comprising the base sequence 5'-GCTCCTCCTCCCTC-3', which also exists, with some differences, in other short reiteration sequences of herpes simplex virus type 1, Epstein-Barr virus, and related human cellular DNA. Furthermore, the tandem repeat array was located in close proximity to the left end of the viral genome and may functionally be involved in viral replication.

Animals↗

PubMed1989

Determination of tissue folate composition by affinity chromatography followed by high-pressure ion pair liquid chromatography.

A recent report from this laboratory described the use of affinity chromatography for the isolation of pure folates from tissue extracts (J. Selhub, B. Darcy-Vrillon, and D. Fell (1988) Anal. Biochem. 168, 247-251). The present study was undertaken to develop chromatographic procedures for quantitative analysis of the individual folates in the affinity-purified mixture. Methods were devised whereby mixtures containing pteroylglutamates (PteGlu1-7) were batch reduced to the dihydro, H2PteGlu1-7, and tetrahydro, H4Pte-Glu1-7, forms. The 5-methylH4PteGlu1-7 and the 10-formylH4PteGlu1-7 series were prepared from H4Pte-Glu1-7. These compounds were used to calibrate a liquid chromatographic system for the resolution of folate mixtures. This system included reverse-phase ion pair chromatography and a diode array detector. A mixture containing oxidized and reduced PteGlu1-7, a total of 35 derivatives, was separated into seven clusters arranged in an order of increasing number of glutamate residues. Each cluster was represented by two or more peaks which were due to folates that differed in the pteridine ring structure but had the same number of glutamate residues. In clusters containing mono and diglutamyl derivatives the 10-formyltetrahydro-, the tetrahydro-, and the dihydrofolate forms appeared as separate peaks while those representing folic acid and 5-methyl-tetrahydrofolate derivatives eluted in coinciding peaks. This hierarchy was maintained in the following clusters except for increasing tendency of the former three forms of folates to elute in the same peak. The number of glutamate residues of any eluting folate can be determined on the basis of retention time in relation to those of the clusters. The pteridine ring structure of that same folate can be determined on the basis of its elution position within that cluster and spectral characteristics determined by the diode array detection system. If that position is common for more than one derivative then identification is based on differential spectral properties. Using uv absorption signals at 280 nm to determine indiscriminate folate activity, absorption signals at 350 nm are used to identify folic acid and dihydrofolate derivatives and signals at 258 nm are used to identify 10-formyltetrahydrofolate derivatives. These principles were incorporated into mathematic expressions which were used for quantitative resolution of simulated mixtures containing oxidized and reduced PteGlu5 and for the analysis of folate composition in rat liver, human milk, and cows milk.

Animals↗

PubMed1989

Anion-exchange high-performance liquid chromatographic determination of ascorbic acid and hexavalent chromium in rat lung preparations after treatment with sodium chromate in vitro and in vivo.

Simultaneous analysis of ascorbic acid and chromium (VI) in soluble fractions and bronchoalveolar lavage fluids of rat lungs treated with sodium chromate in vitro and in vivo was performed by anion-exchange high-performance liquid chromatography coupled to a photodiode-array detector. Absorbances at 265 and 370 nm were used for the determination of ascorbic acid and chromium (VI), respectively. The calibration graphs of standard solutions were linear in the test ranges of ascorbic acid an chromium (VI) (below 10 and 8 ppm, respectively). The detection limits of ascorbic acid and chromium (VI) were 1 and 0.5 ng, respectively. The recovery of ascorbic acid from lung tissues homogenized at pH 7.4 was 99%, and that of chromium (VI) was 96%, when tissues were homogenized under alkaline conditions (pH 11.4). Using this method, ascorbic acid levels in the soluble fractions and lavage fluids of normal rat lungs were determined. In the lung of a rat intratracheally injected with a saline solution of sodium chromate, ascorbic acid decreased to 80% of the normal level, and ca. 90% of the chromium (VI) was reduced within 4 min after injection, indicating that the ascorbic acid-related reduction of chromium (VI) is very rapid. The present method will be useful for studies of the reduction of chromium (VI) by ascorbic acid in biological systems.

Animals↗

PubMed1992

Isolation and expression of linked zinc finger gene clusters on human chromosome 11q.

Proteins that share conserved "zinc finger" motifs represent a class of DNA-binding proteins that have been shown to play a fundamental role in regulating gene expression and to be involved in a number of human hereditary and malignant disease states. We have isolated, characterized, and mapped zinc finger-encoding genes specific to human chromosome 11q to investigate their possible association in the molecular pathogenesis of several disease loci mapped to this chromosome. An arrayed chromosome 11q cosmid library was screened using a degenerate oligonucleotide corresponding to the H/C link consensus sequence of the Drosophila Kruppel zinc finger gene, resulting in the isolation of six putative zinc finger genes. Three of the genes (ZNF123, ZNF125, and ZNF126) were analyzed and shown to contain tandemly repeated zinc finger motifs of the C2-H2 class. All three novel genes were found to be expressed in normal adult human tissues, although the tissue-specific pattern of expression differs markedly. Isolated zinc finger genes were regionally mapped on chromosome 11 using fluorescence in situ suppression hybridization and demonstrated clustering of the genes at 11q13.3-11q13.4 and 11q23.1-11q23.2. Analysis of in situ hybridization to interphase nuclei demonstrated a maximum distance of 1 Mb separating distinct finger genes. This analysis defines two linked multigene families of zinc finger genes to chromosome bands associated with a high frequency of specific translocations associated with malignancies.

Amino Acid Sequence↗

PubMed1990

Optimal excitation of multiapplicator systems for deep regional hyperthermia.

A method is proposed for determining the excitation amplitudes and phases of the elements of electromagnetic multiapplicator systems for optimizing the specific absorption rate (SAR) distribution around a deep-seated tumor. In this method, the ratio of the power dissipated in the tumor to a weighted summation of the powers supplied to the surrounding regions is optimized. The optimization procedure is combined with a recently proposed effective technique for analysis of various electromagnetic scattering and interaction problems. The general principle is applied to a two-dimensional problem of a piecewise homogeneous cylinder heated by an array of electric current filaments placed outside the cylinder. Numerical simulations are performed to check the effectiveness of the approach. The results demonstrate that using this optimization method, improved SAR distributions can be achieved. The extension to three-dimensional configurations is discussed.

Electromagnetic Phenomena↗

PubMed1977

Intercellular junctions in the central nervous system of insects.

The intercellular junctional complexes in the central nervous system (CNS) from a variety of insect species have been examined by thin-sectioning and freeze-fracturing techniques. Of particular concern has been the fine-structural basis of the blood-brain barrier observed to be present in the outer perineurial layer around the avascular insect CNS. The basis of this has been found in the form of tight junctions (zonulae occludentes) present both in sections and in replicas of the perineurium. In the latter, they appear as one or two simple linear ridges, lying parallel to the outer surface, which occasionally display overlapping. The complex geometry of the interdigitating perineurial cells apparently permits such a relatively simple series of ridges to function as a barrier, since tracers are found not to penetrate beyond this level into the underlying nervous tissue. Such evidence is supported by microprobe X-ray analysis of lanthanum-incubated tissues, the perineurium compared with the glia-ensheathed axons showing the presence and absence of lanthanum, respectively. Possible physiological mechanisms that could operate 'in vitro' to maintain the blood-brain barrier are also considered. Other intercellular junctions such as desmosomes, septate junctions and gap junctions are found in the perineurial layer too, the last exhibiting EF particle plaques and PF pits. Glia-glia junctions also occur in some insect species; they include desmosomes, inverted gap junctions and occasional tight junctions. Septate, gap and tight junctions are also found on the membranes of tracheoles penetrating the CNS. Short, ridge-like elaborations and other particle arrays are found on the PF on the axon surfaces and the significance of these structures is discussed.

Animals↗

PubMed1991

Monoclonal antibodies to inner ear antigens: II. Antigens expressed in sensory cell stereocilia.

To develop biological reagents for investigating structure-function relationships in the organ of Corti, we have raised monoclonal antibodies, (MAb) to inner ear tissues. Our first series of antibodies prepared after intrasplenic immunization of mice with guinea pig tissues, identified antigens restricted to supporting cell structures, but no hair cell specific antibodies were developed [Zajic et al., Hear. Res. 52, 59-72, 1991]. In this report we describe the isolation, binding specificity and initial characterization of the stereocilia-binding monoclonal antibodies, KHRI-4, and KHRI-5. Mice were immunized with avian, amphibian and mammalian sensory hair cell-containing tissues and antibodies were screened for selective binding to cochlear extracts in ELISA. In the inner ear, KHRI-4 and KHRI-5 bind specifically to stereocilia in both avian and mammalian cochlear and vestibular tissue preparations using immunofluorescence and immunoperoxidase assays. In other tissues only certain cells of mesothelial origin, such as smooth muscle in gut and the arteriolar vasculature, were stained by KHRI-4 indicating that the antigenic structure defined by this antibody has limited distribution. KHRI-5 binding could be detected in other tissues only at high antibody concentrations suggesting that the gene product identified by this antibody is also weakly expressed in other cell lineages. Western blot analysis showed that KHRI-4 and -5 detect different protein complexes. KHRI-4 identifies an antigenic structure common to gut, cochlea, vestibular tissue and cultured fibroblasts consisting of a approximately 195 and a 230 kDa heterodimer designated p195/230. KHRI-5 binds to a prominent approximately 200-210 kDa band in Western blots of cochlear tissues, gut and fibroblasts. In immunoprecipitation experiments, KHRI-5 precipitated three proteins of Mr approximately 200-210, 230 and 260 kDa indicating that the approximately 200-210 kDa protein carrying the epitope for this antibody is a member of a heterotrimer complex. Our results show that these protein complexes are structural components of stereocilia and that the same proteins are arrayed in conjunction with the actin stress fibers of cultured mesothelial cells. Thus, they are likely to be important for maintaining the actin structure of stereocilia essential to transduction in sensory hair cells.

Animals↗

PubMed2026

Pervasive positive selection on X-linked ampliconic genes in primates.

Mammalian sex chromosomes harbour ampliconic gene families, which are multi-copy genes with ≥97% sequence identity, predominantly expressed in testis tissue and essential for male fertility. The amplification of testis-specific genes is conserved across mammals, yet the specific gene families that expand show striking lineage-specific variation. Previous studies suggest a dynamic turnover with adaptive evolution for several of these families, but their analysis has been limited by the quality of reference genomes of repetitive regions. To characterise the molecular evolutionary processes of ampliconic gene families on both sex chromosomes, we analysed telomere-to-telomere genome assemblies from eight primate species spanning 25 million years of evolution. We identified 53 X-linked and 19 Y-linked ampliconic gene families with dynamic copy number variation. Gene conversion through palindromic pairing and tandem arrays maintained high sequence similarity despite accumulating mutations. X-linked families maintained conserved chromosomal positions despite copy number changes, whereas Y-linked families showed frequent positional turnover. Strikingly, multiple X-linked families (GAGE, SSX, CSAG, and VCX) showed pervasive positive selection across the primate phylogeny and multiple (MAGEB, CT45, HSFX) showed lineage specific positive selection. Y-linked families predominantly evolve under purifying selection. Examining intraspecific copy number variation of the X-linked ampliconic families in chimpanzees, humans, and gorillas, we found variation among individuals but clear differences between species, with the largest families varying the most. These patterns could suggest that sperm competition, meiotic drive, or dosage-dependent selection drive the rapid, lineage-specific evolution of testis-expressed ampliconic genes in primates.

Journal Article↗