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N Lieska

Publications and source records attributed to N Lieska.

At least 19 recordsLinked to original sources
PubMed2001

The ulnar nerve at the elbow and its local branching: an anatomic study.

Thirty nine cadaver elbows were dissected and the branching of the ulnar nerve, as well as the cubital tunnel and adjacent potential sites of nerve compression were studied. An arcade of Struthers was present in 26 specimens and Osborne's ligament was present in all specimens. A discrete flexor pronator aponeurosis overlying the ulnar nerve was present in 17 specimens. An average of one (range, 0-3) capsular nerve branches were noted. These originated an average 7 mm proximal (range, 45 mm proximal to 24 mm distal) to the medial epicondyle. An average of three (range, 1-6) motor branches to the flexor carpi ulnaris muscle were noted, and one of these originated proximal to the medial epicondyle in two specimens. Significant variation was noted in the capsular and motor branching of the ulnar nerve. Care must be taken to identify the motor branches of the ulnar nerve when performing a transposition.

Dissection↗
PubMed1997

A subpopulation of reactive astrocytes at the immediate site of cerebral cortical injury.

We have identified an early-appearing intermediate filament-associated protein (IFAP-70/280 kDa) in radial glia and their immediate derivatives. This IFAP is absent in the adult CNS. In this study, we examined the reexpression of this early glial differentiation trait in rat reactive astrocytes induced by stab injury of the cerebrum. Double-label immunofluorescence microscopy demonstrated that by 36 h postlesion, IFAP-70/280 kDa was present in a few GFAP-positive astrocytes in the area adjacent to the wound. As the gliotic reaction progressed, the number of IFAP-positive reactive astrocytes increased and by 5-6 days postlesion, IFAP-70/280 kDa was present in most of the hypertrophied astrocytes in tissue immediately adjacent to the wound. By 8 days postlesion, while the number of IFAP-negative reactive astrocytes away from the wound diminished, the IFAP-containing reactive astrocytes close to the wound persisted. Concurrently, they began to change from a stellate form to an elongated shape, with their longitudinal axes radiating from the wound. The immunoreactivity of this IFAP started to diminish at 20 days postlesion, and by 30 days postlesion, it was not observed in the remaining gliotic cells. These results demonstrate that reactive astrocytes induced by stab-wound injury can be divided into two subtypes: persistent IFAP-70/280 kDa-containing cells which are close to the wound in the area of the glial scar and transient IFAP-70/280 kDa-negative cells which are farther from the wound. The reappearance of IFAP-70/280 kDa also suggests that some reactive astrocytes have the capacity to recapitulate early developmental stages.

Animals↗
PubMed1996

Immunomicroscopy of neurofilaments in chromaffin cells of the adult bovine adrenal gland.

Neurofilaments (NFs) represent a class of intermediate filaments which are highly specific for neurons. The most abundant of the native NFs is the 68 kD subunit (NF-L). Chromaffin cells of the adrenal medulla express NF subunits under culture conditions. However, NF expression in situ is questionable. It has been reported that as chromaffin cell precursors mature and begin to express catecholamine-synthesizing enzymes, their neuronal traits are extinguished and they become endocrine-like cells. This study reports that while NF-L may be lacking in the adrenal medulla of some species, NF-L immunoreactivity is clearly present in the adult bovine adrenal medulla. Immunofluorescence microscopy of bovine chromaffin cells in culture demonstrated NF immunoreactivity localized to both thin, highly ramified filaments present throughout the cell and frequently to an intensely immunofluorescent spot located near the nucleus. Double-label immunofluorescence microscopy and immunoblot analysis also demonstrated NF-L immunoreactivity in mature chromaffin cells of the bovine adrenal gland. In vitro and in situ immunofluorescence results were confirmed by immunogold and immunoperoxidase labelling, respectively. In both cases, NF-L immunoreactivity was associated with filaments in close proximity to the nucleus. Additionally, a spheroidal aggregate of immunogold-labelled NFs was seen adjacent to the nucleus in cultured cells. In conclusion, NF-L in bovine chromaffin cells demonstrates that at least one neuronal trait persists in these catecholamine-producing cells of the mature adrenal gland. In addition, this study emphasizes the fact that interspecies comparisons must be interpreted with caution, especially when attempting to formulate a generalized hypothesis.

Adrenal Medulla↗
PubMed1994

Immunofluorescence and biochemical studies of the type VI collagen expression by human glioblastoma cells in vitro.

The human glioblastoma cell line U-87 MG was found to express a 140 kD polypeptide which was recognized on immunoblot analysis by a monoclonal antibody to type VI collagen. This polypeptide was digestible by a highly purified bacterial collagenase. After treatment of U-87 MG cells by pepsin, the protein profile revealed the two major pepsin-resistant fragments identical in Mr to those of collagen VI extracted from human placenta. The respective peptide maps from V8 protease one-dimensional gels of these two fragments were identical to those obtained with human collagen VI. Immunofluorescent staining by antibodies to type VI collagen was observed in the extracellular matrix. Moreover, U-87 MG cells were found to be positive for A2B5, a cell surface marker specific for O-2A type glial precursor cells. These data indicate that the human glioblastoma cell line U-87 MG exhibits the properties of glial precursor cells and expresses collagen type VI in vitro. This cell line therefore may prove valuable for comparative investigations of the regulation of type VI collagen synthesis, and may be useful as a model to study the function and pathological importance of type VI collagen in human brain tumours, both in vitro and in vivo.

Biomarkers↗
PubMed1993

Expression of 300-kilodalton intermediate filament-associated protein distinguishes human glioma cells from normal astrocytes.

The availability of biochemical markers to distinguish glioma cells from normal astrocytes would have enormous diagnostic value. Such markers also may be of value in studying the basic biology of human astrocytomas. The vimentin-binding, 300-kDa intermediate filament (IF)-associated protein (IFAP-300kDa) has recently been shown to be developmentally expressed in radial glia of the central nervous system of the rat. It is not detected in the normal or reactive astrocytes of the adult rat nor in neonatal rat brain astrocytes in primary culture. In the present study, double-label immunofluorescence microscopy using antibodies to IFAP-300kDa and glial fibrillary acidic protein (GFAP, an astrocyte-specific IF structural protein) identifies this IFAP in GFAP-containing tumor cells from examples of all three major types of human astrocytomas (i.e., well-differentiated, anaplastic, and glioblastoma multiforme). Astrocytoma cells in primary cultures prepared from all three astrocytomas also express this protein. It is not detectable in normal adult brain tissue. Immunoblot analyses using the IFAP-300kDa antibody confirm the presence of a 300-kDa polypeptide in fresh astrocytoma preparations enriched for IF proteins. These results suggest the utility of IFAP-300kDa as a marker for identification of human glioma cells both in vitro and in situ.

Astrocytes↗
PubMed1993

Immunotyping of radial glia and their glial derivatives during development of the rat spinal cord.

The differentiation of glia in the central nervous system is not well understood. A major problem is the absence of an objective identification system for involved cells, particularly the early-appearing radial glia. The intermediate filament structural proteins vimentin and glial fibrillary acidic protein have been used to define the early and late stages, respectively, of astrocyte development. However, because of the non-specificity of vimentin and the temporal overlap in expression patterns of both proteins, it is difficult to refine our view of the process. This is especially true of the early differentiation events involving radial glia. Using the developmentally-expressed intermediate filament-associated protein IFAP-70/280 kD in conjunction with vimentin and glial fibrillary acidic protein markers, a comprehensive investigation of this problem was undertaken using immunofluorescence microscopy of developing rat spinal cord (E13-P28 plus adult). The phenotypes of the cells were defined on the basis of their immunologic composition with respect to IFAP-70/280 kD (I), vimentin (V) and GFAP (G). A definitive immunotype for radial glia was established, viz, I+/V+/G-; thus reliance upon strictly morphological criteria for this early developmental cell was no longer necessary. Based upon the immunotypes of the cells involved, four major stages of macroglial development were delineated: (1) radial glia (I+/V+/G-); (2) macroglial progenitors (I+/V+/G+); (3) immature macroglia (I-/V+/G+); and (4) mature astrocytes (I-/V+/G+ primarily in white matter and I-/V-/G+, the predominant type in gray matter). It is of interest to note that the cells of the floor plate were distinguished from radial glia by their lack of IFAP-70/280 kD immunoreactivity. Introduction of the IFAP-70/280 kD marker has therefore provided a more refined interpretation of the various differentiation stages from radial glia to mature astrocytes.

Animals↗
PubMed1992

Distinct developmental subtypes of cultured non-stellate rat astrocytes distinguished by a new glial intermediate filament-associated protein.

The nature and tissue origin of cultured non-stellate astrocytes have not been defined. On the basis of immunofluorescence microscopy using multiple double-labeling with antibodies to glial fibrillary acidic protein (GFAP), vimentin, and the recently identified intermediate filament-associated protein (IFAP)-70/280kD, four distinct astrocytic subtypes were definable in neonatal rat brain astrocytes in culture. All of these were of the non-stellate type on the basis of morphology. Similar examination of developing rat cerebral cortex identified these same subtypes as distinct differentiation states of astrocytes. These findings indicate that the parallel developmental events can be studied in vitro.

Animals↗
PubMed1992

Colchicine-sensitive and colchicine-insensitive intermediate filament systems distinguished by a new intermediate filament-associated protein, IFAP-70/280 kD.

A monoclonal antibody was produced, using as antigen a BHK-21 cytoskeletal preparation enriched in intermediate filaments (IF) and their associated proteins. This antibody reacted exclusively with a reproducible set of 70-280 kD polypeptides present in minor quantities in this preparation, as detected by immunoblot analysis. Based upon several criteria, this immunologically related group of polypeptides was designated as IFAP-70/280 kD (IF-Associated Protein): (1) it co-isolated with IF in vitro, (2) it co-localized (by both immunofluorescence and immunoelectron microscopy) with IF in situ in all stages of cell spreading, and (3) it segregated in vitro with the 54/55 kD (desmin/vimentin) structural IF subunit proteins of BHK cells through two cycles of in vitro disassembly/assembly. Immunogold labeling further localized IFAP-70/280 kD to regions of parallel or loosely bundled IF in situ, suggesting a role in regulating the supramolecular organization of IF. When this monoclonal antibody was used for double-label immunofluorescence observations of colchicine-treated BHK cells, it demonstrated the presence of colchicine-sensitive and colchicine-insensitive IF. Anti-IFAP-70/280 kD localized entirely to the drug-induced juxtanuclear IF cap, while a polyclonal antibody directed against the desmin/vimentin structural IF subunits and the previously characterized monoclonal anti-IFAP-300 kD [Yang et al., 1985; J. Cell Biol. 100:620] localized to both the juxtanuclear IF cap and a colchicine-insensitive IF network peripheral to the cap in the same cells. The colchicine-insensitive IF pattern often exhibited similarities to that observed for the actin-based stress fiber system, suggesting that stress fiber association may be an additional factor in IF organization.

Animals↗
PubMed1991

Reconstitution of the filamentous backbone of lens beaded-chain filaments from a purified 49kD polypeptide.

The beaded-chain filaments unique to the fiber cells of the crystalline lens are composed of a linear array of spheroidal particles which appear to be connected by a filamentous backbone. In order to determine the existence of the putative backbone and to characterize its constituents, one of the major proteins associated with beaded-chains in the chicken lens was investigated. 49kD was isolated in an enriched fraction derived from the 8M urea extract of the lens cell water-insoluble residue. The polypeptide (which exists in several charge isoforms, the major at pI 5.2) was purified sequentially by gel filtration on Sephacryl S-200, hydrophobic interaction chromatography on phenyl-Sepharose, and anionic exchange chromatography on Mono Q, all under denaturing conditions. Immunoblot analyses established that 49kD was immunologically distinct from vimentin, actin, and tubulin/MAPs (representing the three classes of cytoplasmic filaments), as well as from the crystallins. Amino acid analyses demonstrated compositional differences for 49kD compared with lens actin and vimentin, and one- and two-dimensional peptide mapping of 49kD and vimentin revealed no homology. Electron microscopy demonstrated that short, contorted filaments were produced upon removal of purified 49kD from urea to low-salt buffers. In the presence of physiological salt concentrations 49kD assembled into extensive 4-6nm diameter, straight filaments similar to the backbone seen in native beaded-chain filaments, but morphologically distinct from the other cytoplasmic filament classes.

Amino Acids↗
PubMed1991

Expression and distribution of cytoskeletal IFAP-300kD as an index of lens cell differentiation.

By their implication in the organization of the intermediate filament (IF) cytoskeleton, IF-associated proteins (IFAPs) can delineate subsets of the same IF type within a cell; moreover, they are proving useful as markers of the differentiation states of certain cells. For these reasons the expression of the vimentin-associated IFAP-300kD was investigated in the constantly differentiating cell lineage of the adult bovine lens. Immunofluorescence microscopy and immunoblot analysis were employed using a monoclonal anti-IFAP-300kD and a rabbit anti-lens vimentin. Cultures of adult lens epithelial cells were immunopositive for the IFAP. By double-label studies the IFAP-300kD pattern co-localized with that of the vimentin-type IF; moreover, the IFAP pattern co-distributed with that of both colchicine-sensitive and -insensitive IF systems. IFAP-300kD was also present in a co-distributing pattern with vimentin IF in fresh lens epithelial cells on whole mounts. There was a differential expression of the IFAP in the lens fiber cells in that those of the cortex exhibited the IFAP and vimentin IF, while both proteins were absent from the nuclear fiber cells. Furthermore, there was a differential distribution of the IFAP within the cortical fiber cells in that the IFAP localized only with a paramembranal subset of IF. Immunoblot analysis supported the presence of IFAP-300kD in the lens cytoskeletal fraction. IFAP-300kD thus identified a subset of vimentin IF whose location may have functional significance for the cortical fiber cell. The changes in the IFAP's expression and distribution pattern throughout lens cell differentiation in the adult organ suggest the usefulness of IFAP-300kD as a potential marker in studying lens cell differentiation in vitro.

Animals↗
PubMed1986

The amino acid sequence of human cardiac troponin-C.

The amino acid sequence of human cardiac troponin-C (TN-C) has been rabbit cardiac TN-C. These results show that troponin-C is one of the Edman degradation. The sequence, which consists of 161 amino acid residues, is identical to that of the ox heart and as such is capable of binding three calcium ions. It differs at only one position from that of rabbit cardiac TN-C. These results show that troponin C is one of the most highly conserved proteins known.

Amino Acid Sequence↗
PubMed1985

A 300,000-mol-wt intermediate filament-associated protein in baby hamster kidney (BHK-21) cells.

Native intermediate filament (IF) preparations from the baby hamster kidney fibroblastic cell line (BHK-21) contain a number of minor polypeptides in addition to the IF structural subunit proteins desmin, a 54,000-mol-wt protein, and vimentin, a 55,000-mol-wt protein. A monoclonal antibody was produced that reached exclusively with a high molecular weight (300,000) protein representative of these minor proteins. Immunological methods and comparative peptide mapping techniques demonstrated that the 300,000-mol-wt species was biochemically distinct from the 54,000- and 55,000-mol-wt proteins. Double-label immunofluorescence observations on spread BHK cells using this monoclonal antibody and a rabbit polyclonal antibody directed against the 54,000- and 55,000-mol-wt proteins showed that the 300,000-mol-wt species co-distributed with IF in a fibrous pattern. In cells treated with colchicine or those in the early stages of spreading, double-labeling with these antibodies revealed the co-existence of the respective antigens in the juxtanuclear cap of IF that is characteristic of cells in these physiological states. After colchicine removal, or in the late stages of cell spreading, the 300,00-mol-wt species and the IF subunits redistributed to their normal, highly coincident cytoplasmic patterns. Ultrastructural localization by the immunogold technique using the monoclonal antibody supported the light microscopic findings in that the 300,000-mol-wt species was associated with IF in the several physiological and morphological cell states investigated. The gold particle pattern was less intimately associated with IF than that defined by anti-54/55 and was one of non-uniform distribution along IF, being clustered primarily at points of proximity between IF, where an amorphous, proteinaceous material was often the labeled element. Occasionally, "bridges" of label were seen extending outward from such clusters on IF. Gold particles were infrequently bound to microtubules, microfilaments, or other cellular organelles, and when so, IF were usually contiguous. During multiple cycles of in vitro disassembly/assembly of the IF from native preparations, the 300,000-mol-wt protein remained in the fraction containing the 54,000- and 55,000-mol-wt structural subunits, whether the latter were in the soluble state or pelleted as formed filaments. In keeping with the nomenclature developed for the microtubule-associated proteins (MAPs), the acronym IFAP-300K (intermediate filament associated protein) is proposed for this molecule.

Animals↗
PubMed1985

Purification of the 300K intermediate filament-associated protein and its in vitro recombination with intermediate filaments.

IFAP-300K is a 300,000-mol-wt intermediate filament-associated protein previously identified in the baby hamster kidney fibroblastic cell line (BHK-21) by a monoclonal antibody (Yang H.-Y., N. Lieska, A. E. Goldman, and R. D. Goldman, 1985, J. Cell Biol., 100: 620-631). In the present study, this molecule was purified from the high salt/detergent-insoluble cytoskeletal preparation of these cells. Gel filtration on Sephacryl S-400 in the presence of 7.2 M urea allowed separation of the high molecular weight fraction from the structural intermediate filament (IF) subunits desmin and vimentin, designated 54K and 55K, respectively, and other low molecular weight polypeptides. DE-52 cellulose chromatography of the high molecular weight fraction using a linear NaCl gradient in 8 M urea yielded a pure 300,000-mol-wt species which was confirmed to be IFAP-300K by immunological and peptide mapping criteria. Two-dimensional PAGE of native BHK IF preparations followed by immunoblot analysis demonstrated the inability of the IFAP-300K-immunoreactive material to enter the first dimensional gel except as a 200,000-mol-wt doublet which presumably represented a major proteolytic derivative of IFAP-300K. The molecule's pl of 5.35, as determined by chromatofocusing, and its amino acid composition were extremely similar to those of BHK cell vimentin/desmin despite their non-identity. Ultrastructurally, IFAP-300K preparations in low salt buffers existed as particles composed of one or two elliptical units measuring 16 X 20 nm. In physiological salt buffers, the predominant entities were large, elongated aggregates of the elliptical units, which were able to be decorated by using the immunogold technique with monoclonal anti-IFAP-300K. Compared with the morphology of homopolymer vimentin IF, in vitro recombination studies using column-purified vimentin and IFAP-300K demonstrated the additional presence of aggregates similar in appearance to IFAP-300K at points of contact between IFs. Antibody decoration and immunogold labeling of these recombined preparations using rabbit antidesmin/vimentin and monoclonal anti-IFAP-300K confirmed the identity of the inter-filament, amorphous material as IFAP-300K. The presence of IFAP-300K at many points of intersection and lateral contact between IFs, as well as at apparent inter-filament "bridges," in these recombined specimens was identical to that seen both in situ and in native IF preparations. No such co-sedimentation was found in vitro between actin and IFAP-300K. No effects of IFAP-300K upon the kinetics of IF polymerization were detected by turbidimetric measurements.

Amino Acids↗
PubMed1983

Lens actin: purification and localization.

Actin was purified from the chick lens using DEAE-52 column chromatography followed by hydroxylapatite chromatography. The antibody produced against the purified actin cross-reacted specifically with lens actin from other species in addition to smooth and skeletal muscle actin and labelled the stress bundles of cultured fibroblasts. Actin was localized, using immunological methods, primarily to the plasma membrane of the epithelial and fiber cells of the chick and human lens. Actin filaments were also identified by HMM S-1 labeling in bovine cortical fiber cells. Using this procedure, the actin filaments were found throughout the fiber cell but were mainly concentrated near the plasma membrane and in cell processes. They formed a population distinct from the beaded filaments. The initial DEAE-52 column chromatography was also useful in the initial purification of lens fiber cell intermediate filament protein and two species of beta-crystallins.

Actins↗
PubMed1981

Electron microscopy supports a fibrous substructure for lens intermediate filaments.

The substructure of intermediate filaments from bovine lens cortical fiber cells was investigated by electron microscopy. Native filaments and synthetic ones regenerated from the total cytoskeletal extract and from the three purified subunits were examined. The morphologies from these various sources were essentially identical, with the exception that filaments reconstituted from one of the purified polypeptides were much shorter, very contorted and showed strings of aggregated protein. The solid cylindrical, unbranching filaments consisted of a helical arrangement of at least two, 5 nm diameter strands. The evidence indicated that each strand was composed of two, 2 nm diameter protofilaments which were also helically constructed (right-handed) with a periodicity of 11.6 nm. Intermediate filament diameter varied widely (8-14.8 nm, average 11.3 nm) and in a direct, linear manner relative to the apparent progression (helical) angle of the strands across the filaments face. These conclusions were obtained from observations on negatively stained intact filaments and reconstituted 4.4 nm fibrils and on positively stained transverse sections of fixed and embedded filaments.

Animals↗