scholarly journals Distribution of the large aggregating proteoglycan versican in adult human tissues.

1996 ◽  
Vol 44 (4) ◽  
pp. 303-312 ◽  
Author(s):  
B Bode-Lesniewska ◽  
M T Dours-Zimmermann ◽  
B F Odermatt ◽  
J Briner ◽  
P U Heitz ◽  
...  
Keyword(s):  
Core Protein ◽  
Splice Variants ◽  
Polyclonal Antibodies ◽  
Elastic Fiber ◽  
Basal Layer ◽  
Human Tissues ◽  
Connective Tissues ◽  
Fiber Network ◽  
Muscle Tissues ◽  
The Media

We studied the distribution of the large hyaluronan-binding proteoglycan versican (also known as PG-M) in human adult tissues using affinity-purified polyclonal antibodies that recognize the core protein of the prominent versican splice variants VO and V1. Versican was present in the loose connective tissues of various organs and was often associated with the elastic fiber network. Furthermore, it was localized in most smooth muscle tissues and in fibrous and elastic cartilage. Versican staining was also noted in the central and peripheral nervous system, in the basal layer of the epidermis, and on the luminal surface of some glandular epithelia. In blood vessels, versican was present in all three wall layers of veins and elastic arteries. In muscular arteries the immunoreactivity was normally restricted to the tunica adventitia. However, it appeared in the media and the split elastica interna of atherosclerotically transformed vessel walls. Our survey of the distribution of versican in normal human tissues now forms the basis for extended studies of potentially aberrant versican expression during pathogenic processes.

Synaptonemal complex proteins: occurrence, epitope mapping and chromosome disjunction

1994 ◽  
Vol 107 (10) ◽  
pp. 2749-2760 ◽  
Author(s):  
M.J. Dobson ◽  
R.E. Pearlman ◽  
A. Karaiskakis ◽  
B. Spyropoulos ◽  
P.B. Moens
Keyword(s):  
Synaptonemal Complex ◽  
Fusion Proteins ◽  
Epitope Mapping ◽  
Core Protein ◽  
Binding Capacity ◽  
Polyclonal Antibodies ◽  
Meiotic Chromosome ◽  
Immunogold Labelling ◽  
Chromosome Disjunction ◽  
Metaphase I

We have used polyclonal antibodies against fusion proteins produced from cDNA fragments of a meiotic chromosome core protein, Cor1, and a protein present only in the synapsed portions of the cores, Syn1, to detect the occurrence and the locations of these proteins in rodent meiotic prophase chromosomes. The 234 amino acid Cor1 protein is present in early unpaired cores, in the lateral domains of the synaptonemal complex and in the chromosome cores when they separate at diplotene. A novel observation showed the presence of Cor1 axial to the metaphase I chromosomes and substantial amounts of Cor1 in association with pairs of sister centromeres. The centromere-associated Cor1 protein becomes dissociated from the centromeres at anaphase II and it is not found in mitotic metaphase centromeres. The extended presence of Cor1 suggests that it may have a role in chromosome disjunction by fastening chiasmata at metaphase I and by joining sister kinetochores, which ensures co-segregation at anaphase I. Two-colour immunofluorescence of Cor1 and Syn1 demonstrates that synapsis between homologous cores is initiated at few sites but advances rapidly relative to the establishment of new initiation sites. If the rapid advance of synapsis deters additional initiation sites between pairs of homologues, it may provide a mechanism for positive recombination interference. Immunogold epitope mapping of antibodies to four Syn1 fusion proteins places the amino terminus of Syn1 towards the centre of the synaptonemal complex while the carboxyl terminus extends well into the lateral domain of the synaptonemal complex. The Syn1 fusion proteins have a non-specific DNA binding capacity. Immunogold labelling of Cor1 antigens indicates that the lateral domain of the synaptonemal complex is about twice as wide as the apparent width of lateral elements when stained with electron-dense metal ions. Electron microscopy of shadow-cast surface-spread SCs confirms the greater width of the lateral domain. The implication of these dimensions is that the proteins that comprise the synaptic domain overlap with the protein constituents of the lateral domains of the synaptonemal complex more than was apparent from earlier observations. This arrangement suggests that direct interactions might be expected between some of the synaptonemal complex proteins.

scholarly journals Complex CSF3R Protein Isoform Interactions Dictate Cytokine Responsiveness

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 35-35
Author(s):  
Sara L. Seegers ◽  
Amanda Lance ◽  
Lawrence J Druhan ◽  
Belinda R Avalos
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Splice Variant ◽  
Splice Variants ◽  
Polyclonal Antibodies ◽  
Human Cells ◽  
Dependent Manner ◽  
Epitope Tag ◽  
Proliferative Signaling ◽  
Primary Human Cells

CSF3R, the receptors for granulocyte colony stimulating factor, is a critical regulator of neutrophil production. Multiple CSF3R mRNA transcripts have been identified and are annotated in Genbank. The expression and function of the different CSF3R proteins have not been fully elucidated. We generated antibodies specific for two of the identified and annotated isoforms, V3 and V4. CSF3R-V4 is a truncated variant of V1 with a unique C-terminal 34 amino acids and this variant confers enhanced growth signals. Changes in the ratio of V1:V4 isoforms have been implicated in chemotherapy resistance and relapse of AML. CSF3R-V3 is a variant of V1 with a 27 amino acid insertion between two conserved domains in the cytoplasmic portion of the receptor involved in JAK/STAT activation, termed the box 1 and box 2. CSF3R-V3 produces reduced proliferative signaling in response to G-CSF. When V3 is co-expressed with V1, proliferative signaling is reduced in a concentration dependent manner. In order to generate custom rabbit polyclonal antibodies specific for CSF3R-V3 and CSF3R-V4 we used either a peptide that corresponds to a unique amino acid sequence present only in CSF3R-V3 or a peptide specific for a portion of the C-terminal amino acid sequence unique to the CSF3R-V4 isoform conjugated to an immunogenic carrier protein. These immunogens both produced robust immune responses, and the polyclonal antibodies were subsequently purified from bulk sera. Immunoblot analysis of lysates from Ba/F3 cells expressing CSF3R-V1 (V1), CSF3R-V3 (V3), or CSF3R-V4 (V4) demonstrated that both the custom generated anti-CSF3R-V3 and anti-CSF3R-V4 antibodies were very specific, recognizing only the appropriate CSF3R receptor isoform. All three CSF3R splice variants are recognized by commercially available anti-CSF3R (clone LMM741 to CD114), while the anti-CSF3R-V4 custom antibody and the custom anti-CSF3R-V3 antibody recognizes only the CSF3R-V4 and CSF3R-V3 isoforms, respectively. We next sought to detect the CSF3R receptor isoforms in primary human cells. Using our custom antibodies, we detected for the first time, both the CSF3R-V3 and CSF3R-V4 receptor forms in primary neutrophils isolated from healthy donors. Each of the CSF3R isoforms produce unique signaling, and we hypothesized that the observed differences in G-CSF-dependent signaling is produced by the expression level of each receptor isoform via both homodimerization and by heterodimerization of the receptor splice variant proteins. To investigate the potential for heterodimerization of the CSF3R-V1 with the V3 and V4 isoforms, we generated a CSF3R-V1 with a c-terminal epitope tag and co-expressed this construct with both CSF3R-V3 or CSF3R-V4. Immunoprecipitation with an antibody to the epitope tag (recognizing the V1 variant) followed by immunoblotting with the custom anti-V3 or anti-V4 antibodies demonstrated that both CSF3R-V3 and CSF3R-V4 co-immunoprecipitated with CSF3R-V1, in agreement with our hypothesis that the splice variants form receptor heterodimers. Of note, the CSF3R receptor heterodimers are detected even in the absence of G-CSF, thus demonstrating that CSF3R exist as a preformed receptor dimer in an inactive state. In conclusion, we have generated antibodies that specifically detect the CSF3R-V3 and the CSF3R-V4 receptor proteins. These are the first studies to demonstrate the expression of the CSF3R splice variants at the protein level, in both cell lines and primary human cells. In addition, these are the first studies to demonstrate the formation of heterodimers of the CSF3R splice variants, providing a mechanism for the observed alteration in ligand-dependent signaling produced under conditions of altered splice variant expression. Disclosures Avalos: Juno: Membership on an entity's Board of Directors or advisory committees; Best Practice-Br Med J: Patents & Royalties: receives royalties from a coauthored article on evaluation of neutropenia.

Obtaining of Collagen Extracts Used as Biomaterials with Applications in the Medical Field

2020 ◽  
Vol 3 (2) ◽  
pp. 84
Author(s):  
Melat Cherim ◽  
Rodica Sîrbu
Keyword(s):  
Core Protein ◽  
Morphological Structure ◽  
Molecular Structures ◽  
Supramolecular Organization ◽  
Connective Tissues ◽  
Medical Field ◽  
Genetic Types ◽  
Nano Structures ◽  
Different Types ◽  
And Function

Collagen is the core protein of connective tissues: skin, bone, tendon, base membrane, etc. Collagen is actually a family of several different genetic types. Currently they are known, in vertebrates, at least 27 different types of collagens, which shows a remarkable diversity in molecular and supramolecular organization of the tissue distribution and function, discovered and developed over 45 years. They were studied, in detail, 12 main types. Collagen-based bioproducts can be produced in a variety of molecular structures (micro and nano structures) in powder form, hydrogels and injectable solutions, films, membranes and matrices, etc. This paper presents the drying processes that are selected depending on the nature of the extract (undenatured or denatured) and morphological structure bioproduct or sponge, fibers or membranes. The most frequently used procedures for drying are freeze-drying and free drying at a temperature of approx. 25 ° C. Both processes produce no distorts to the extracts They are presented bioproducts derived from collagen which are used in medicine.

Ultrastructural Evidence for Proteoglycans Mediating an Attachment of Type VI Collagen to Banded Collagen Fibrils

1997 ◽  
Vol 3 (S2) ◽  
pp. 153-154
Author(s):  
Douglas R. Keene ◽  
Catherine C. Ridgway ◽  
Renato V. Iozzo
Keyword(s):  
Dermatan Sulfate ◽  
Core Protein ◽  
Solid Phase ◽  
Collagen Fibrils ◽  
Binding Assays ◽  
Connective Tissues ◽  
Type Vi Collagen ◽  
Dermatan Sulfate Proteoglycan ◽  
Individual Type ◽  
Solid Phase Binding

Immunolocalizaton studies of type VI collagen in skin have previously demonstrated that type VI collagen forms a flexible network that anchors large interstitial structures such as nerves, blood vessels, and collagen fibers into the surrounding connective tissues matrix. The purpose of this study is to determine if individual type VI collagen microfilaments might be connected to banded collagen fibrils, thereby stabilizing the network.Solid phase binding assays suggest a specific, high affinity interaction between the core protein of the dermatan sulfate proteoglycan decorin and type VI collagen, and immunocytochemical studies in fetal and neonate rabbit cornea suggest an association of decorin with type VI microfilaments. Other studies in skin and perichondrium have localized decorin to a region between the d and e bands of banded collagen fibrils. However, no direct documentation has demonstrated a specific structural interaction between type VI microfilaments and banded collagen fibrils. We, therefore, sought to determine if type VI microfilaments cross banded collagen fibrils between the “d” and “e” bands.

Novel splice variants of the amyotrophic lateral sclerosis-associated gene VAPB expressed in human tissues

2010 ◽  
Vol 394 (3) ◽  
pp. 703-708 ◽  
Author(s):  
T. Nachreiner ◽  
M. Esser ◽  
V. Tenten ◽  
D. Troost ◽  
J. Weis ◽  
...  
Keyword(s):  
Amyotrophic Lateral Sclerosis ◽  
Splice Variants ◽  
Human Tissues ◽  
Lateral Sclerosis

scholarly journals Structure and expression of fibrillin-2, a novel microfibrillar component preferentially located in elastic matrices

1994 ◽  
Vol 124 (5) ◽  
pp. 855-863 ◽  
Author(s):  
H Zhang ◽  
SD Apfelroth ◽  
W Hu ◽  
EC Davis ◽  
C Sanguineti ◽  
...  
Keyword(s):  
Gene Product ◽  
Elastic Fiber ◽  
Extracellular Proteins ◽  
Connective Tissues ◽  
New Family ◽  
Fibrillin 1 ◽  
Partial Cdna ◽  
Point Determination ◽  
Human Chromosome 5

During the previous cloning of the fibrillin gene (FBN1), we isolated a partial cDNA coding for a fibrillin-like peptide and mapped the corresponding gene (FBN2) to human chromosome 5. (Lee, B., M. Godfrey, E. Vitale, H. Hori, M. G. Mattei, M. Sarfarazi, P. Tsipouras, F. Ramirez, and D. W. Hollister. 1991. Nature [Lond.]. 352:330-334). The study left, however, unresolved whether or not the FBN2 gene product is an extracellular component structurally related to fibrillin. Work presented in this report clarifies this important point. Determination of the entire primary structure of the FBN2 gene product demonstrated that this polypeptide is highly homologous to fibrillin. Immunoelectron microscopy localized both fibrillin proteins to elastin-associated extracellular microfibrils. Finally, immunohistochemistry revealed that the fibrillins co-distribute in elastic and non-elastic connective tissues of the developing embryo, with preferential accumulation of the FBN2 gene product in elastic fiber-rich matrices. These results support the original hypothesis that the fibrillins may have distinct but related functions in the formation and maintenance of extracellular microfibrils. Accordingly, we propose to classify the FBN1 and FBN2 gene products as a new family of extracellular proteins and to name its members fibrillin-1 and fibrillin-2, respectively.

scholarly journals Immunohistochemical localization of glycosaminoglycans and proteoglycans in predentin and dentin of rat incisors.

1990 ◽  
Vol 38 (3) ◽  
pp. 319-324 ◽  
Author(s):  
M Takagi ◽  
H Hishikawa ◽  
Y Hosokawa ◽  
A Kagami ◽  
F Rahemtulla
Keyword(s):  
Dermatan Sulfate ◽  
Core Protein ◽  
Polyclonal Antibodies ◽  
Keratan Sulfate ◽  
Immunohistochemical Localization ◽  
Immunoperoxidase Staining ◽  
Weak Staining ◽  
Dentin Matrix ◽  
Pre Treatment ◽  
Definite Difference

We examined immunocytochemically the type and distribution of glycosaminoglycans and proteoglycans (PG) in predentin and dentin demineralized with EDTA after aldehyde fixation of rat incisors using (a) four monoclonal antibodies (1-B-5,9-A-2,3-B-3, and 5-D-4) which recognize epitopes in unsulfated chondroitin (C0-S), chondroitin 4-sulfate (C4-S), chondroitin 6-sulfate (C6-S), and keratan sulfate (KS) associated with the PG, and (b) monoclonal (5-D-5) and polyclonal antibodies specific for the core protein of large and small dermatan sulfate (DS) PG. Light microscope immunoperoxidase staining after pre-treatment of tissue sections with chondroitinase ABC localized the majority of stainable PG (C4-S, KS, DSPG, C0-S, and C6-S) in predentin and, to a lesser extent (C4-S and small DSPG), in the dentin matrix. The former site demonstrated relatively homogeneous PG distribution, whereas the latter site revealed that strong staining of C4-S and small DSPG was confined mostly to dentinal tubules surrounding odontoblastic processes, with only weak staining in the rest of the dentin matrix. These results indicate that there is not only a definite difference between PG of predentin and dentin but also a selective decrease in the concentration or alteration of these macromolecules during dentinogenesis and mineralization.

scholarly journals Elastic Fibers and Large Artery Mechanics in Animal Models of Development and Disease

2018 ◽  
Vol 140 (2) ◽  
Author(s):  
Maria Gabriela Espinosa ◽  
Marius Catalin Staiculescu ◽  
Jungsil Kim ◽  
Eric Marin ◽  
Jessica E. Wagenseil
Keyword(s):  
Animal Models ◽  
Mechanical Behavior ◽  
Computational Models ◽  
Mechanical Response ◽  
Elastic Fiber ◽  
High Stress ◽  
Elastic Fibers ◽  
Large Artery ◽  
Fiber Network ◽  
Pulsatile Pressure

Development of a closed circulatory system requires that large arteries adapt to the mechanical demands of high, pulsatile pressure. Elastin and collagen uniquely address these design criteria in the low and high stress regimes, resulting in a nonlinear mechanical response. Elastin is the core component of elastic fibers, which provide the artery wall with energy storage and recoil. The integrity of the elastic fiber network is affected by component insufficiency or disorganization, leading to an array of vascular pathologies and compromised mechanical behavior. In this review, we discuss how elastic fibers are formed and how they adapt in development and disease. We discuss elastic fiber contributions to arterial mechanical behavior and remodeling. We primarily present data from mouse models with elastic fiber deficiencies, but suggest that alternate small animal models may have unique experimental advantages and the potential to provide new insights. Advanced ultrastructural and biomechanical data are constantly being used to update computational models of arterial mechanics. We discuss the progression from early phenomenological models to microstructurally motivated strain energy functions for both collagen and elastic fiber networks. Although many current models individually account for arterial adaptation, complex geometries, and fluid–solid interactions (FSIs), future models will need to include an even greater number of factors and interactions in the complex system. Among these factors, we identify the need to revisit the role of time dependence and axial growth and remodeling in large artery mechanics, especially in cardiovascular diseases that affect the mechanical integrity of the elastic fibers.

Characterization of Dendritic Cell-Specific Genes p275 and p306.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 3842-3842
Author(s):  
Ingo Hilgendorf ◽  
Daniel Kurz ◽  
Anita Bringmann ◽  
Lothar Kanz ◽  
Frank Grünebach ◽  
...  
Keyword(s):  
Peripheral Blood ◽  
Splice Variant ◽  
Splice Variants ◽  
Polyclonal Antibodies ◽  
Antigen Presenting Cells ◽  
Poly I:C ◽  
Blood Monocytes ◽  
Splice Form ◽  
Antigen Presenting

Abstract Dendritic cells play an inimitable role in the functioning immune system as they are the most potent antigen presenting cells being able to prime naive T-cells. Their characteristic properties that enable them to take up antigens and present them to leukocytes are due to an expression of specific genes and thus specific proteins that are unique to this subset of antigen-presenting cells. Using a substractive cDNA library based on suppression hybridization between DC cDNA and the reference monocyte cDNA, we identified in DC two differentially expressed genes p275 and p306. p275 codes for a membrane protein and represents a splice isoform of the transport protein NAT-1. The predicted structure of protein p306 is globular, suggesting that the protein is either intracellular or secreted. The expression of both genes was confirmed by RT-PCR using cDNA isolated from peripheral blood monocytes and DC, generated in vitro from monocytes or CD34+ progenitor cells. To further analyze the protein expression polyclonal antibodies were generated by immunization with synthetic peptides deduced from the identified sequences. Interestingly, inhibition of DC differentiation using IL-10 or STI571 (Imatinib) resulted in an impaired expression of both proteins. Utilizing specific primers for two recently described splice variants of p306 we identified a new splice form expressed in DC. While the gene of p306 contains eight exons, splice variant 1 consists of the exons 1,2,4,5,6, and 7 and splice variant 2 contains the exons 1,2,3,4,5,6, a shortened exon 7, and exon 8. The new identified splice form includes the exons 1–7. However, as the open reading frame starts in exon 4, the expressed protein is identical with the one corresponding to splice variant 1. Analyzing different DC populations in peripheral blood we show that p306 is expressed in plasmacytoid, but not myeloid DC. Interestingly, the activation of DC with Toll-like receptor ligands (TLRL) Pam3Cys (TLR2L), Poly I:C (TLR3L), LPS (TLR4L) and R848 (TLR7L) has no influence on the expression of p306. Although the functions of p275 and p306 in DC have yet to be determined, both genes play a role in DC differentiation and are found in different hematopoietic cell populations. Especially p306 might be an interesting marker of plasmacytoid DC as the predicted protein does not resemble any known protein structure.

Binders in Paintings

MRS Bulletin ◽  
1996 ◽  
Vol 21 (12) ◽  
pp. 24-31
Author(s):  
Richard Newman
Keyword(s):  
Thin Layers ◽  
Animal Glue ◽  
Variable Properties ◽  
Connective Tissues ◽  
Ancient Egyptian ◽  
Naturally Occurring ◽  
Synthetic Materials ◽  
The Media ◽  
Japanese Painting ◽  
Adhesive Materials

Many naturally occurring adhesive materials have been used throughout history to bind pigments in paintings. A number of synthetic materials have been added to these during the twentieth century.Availability and tradition can influence the choice of binders made by artists. Probably the most widely used medium throughout history, animal glue, is also the most easily obtained. Glues made from the connective tissues or skins of local animals were major media in ancient Egyptian painting and in Chinese and Japanese painting, as well as in many other cultures throughout world history. In many cases, a variety of natural binders were available, and additional factors influenced the choice of binder by a culture. Different media have highly variable properties that affect how they are used in painting. Among these properties are solubility, the transparency or depth of color that is obtainable with a given pigment, and handling properties–how the paint flows, how quickly it dries, whether it can be applied in very thick and very thin layers, etc. Knowledge of the media utilized in paintings can help us understand the intentions of artists. Medieval European painting can be used as an example.

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