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Characteristic expression of Lewis-antigenic glycolipids in human ovarian carcinoma-derived cells with anticancer drug-resistance.

By comparing ovarian carcinoma-derived KF28 cells with the corresponding anticancer drug-resistant cells, the taxol- and cisplatin-resistant properties were found to be closely related with MDR1 and BSEP, and MRP2 transporters, respectively. In addition to the transporters expression, the amounts of glycolipids, particularly their longer carbohydrate structures, in the resistant cells increased to 3-4-fold of those in the sensitive cells due to enhanced transcription of the respective glycosyltransferases. The major glycolipids in the sensitive and resistant cells were GlcCer and Gb(3)Cer, respectively, and extension of the carbohydrate structure into Lewis antigen characteristically occurred in the resistant cells. Le(b), which was not detected in the cisplatin-resistant cells, was present in the taxol-resistant cells, while Le(x) was present in the cisplatin-resistant cells at a higher concentration than in the taxol-resistant cells. 2-Hydroxy fatty acids were significantly abundant in glycolipids of the resistant cells, but they were not detected in free ceramides or sphingomyelin, indicating that the enhanced synthesis of glycolipids in the resistant cells was not linked with the removal pathway for virulent ceramides derived from sphingomyelin. The resistant cells with abundant glycolipids exhibited lower membrane fluidity than the KF28 cells, and this property might be involved in the anticancer drug-resistance.

Cisplatin↗

Further studies on cell adhesion based on Le(x)-Le(x) interaction, with new approaches: embryoglycan aggregation of F9 teratocarcinoma cells, and adhesion of various tumour cells based on Le(x) expression.

We previously proposed specific interaction of Le(x) (Gal beta 1-->4 [Fuc alpha 1-->3]-GlcNAc beta 1-->3Gal) with Le(x) as a basis of cell adhesion in pre-implantation embryos and in aggregation of F9 teratocarcinoma cells, based on several lines of evidence (Eggens et al., J. Biol Chem (1989) 264:9476-9484). We now present additional evidence for this concept, based on autoaggregation studies of plastic beads coated with glycosphingolipids (GSLs) bearing Le(x) or other epitopes, and affinity chromatography on Le(x)-columns of multivalent lactofucopentaose III (Le(x) oligosaccharide) conjugated with lysyllysine. Comparative adhesion studies of Le(x)-expressing tumour cells vs their Le(x)-non-expressing variants showed that only Le(x)-expressing cells adhere to Le(x)-coated plates and are involved in tumour cell aggregation, in analogy to F9 cell aggregation. The major carrier of Le(x) determinant in F9 cells is not GSL but rather polylactosaminoglycan ('embryoglycan'), and we demonstrated autoaggregation of purified embryoglycan in the presence of Ca2+, and reversible dissociation in the absence of Ca2+ (addition of EDTA). Defucosylated embryoglycan did not show autoaggregation under the same conditions. Thus, Le(x)-Le(x) interaction has been demonstrated on a lactosaminoglycan basis as well as a GSL basis. A molecular model of Le(x)-Le(x) interaction based on minimum energy conformation with involvement of Ca2+ is presented.

Carbohydrate Sequence↗

Acquisition of P-selectin binding activity by en bloc transfer of sulfo Le(x) trisaccharide to the cell surface: comparison to a sialyl Le(x) tetrasaccharide transferred on the cell surface.

Sialyl Le(x), NeuNAcalpha2 --> 3Galbeta1 --> 4(Fucalpha1 --> 3)GlcNAcbeta --> R, is known to be a ligand for E-selectin in various assays. The sulfated counterpart of sialyl Le(x), sulfo Le(x), (Sulfo --> 3) Galbeta1 --> 4 (Fucalpha1 --> 3) GlcNAcbeta --> R, was also shown to be a ligand for E-selectin in solid-phase assays employing immobilized oligosaccharides. In order to determine whether sulfo Le(x) structure on the cell surface also works as E-selectin or P-selectin ligand, a novel approach for in vitro transfer of oligosaccharides (S. Tsuboi, Y. Isogai, N. Hada, J. K. King, O. Hindsgaul, and M. Fukuda (1996) J. Biol. Chem. 271, 27213-27216) was utilized. A synthetic GDP-fucose harboring sialyl Le(x) or sulfo Le(x) oligosaccharide was enzymatically transferred to Chinese hamster ovary (CHO) cells with a milk fucosyltransferase. The resultant cells, CHO-sialyl Le(x) and CHO-sulfo Le(x) were tested for adhesion to E-selectin. IgG or P-selectin. IgG chimeric protein coated on plates. The results indicate that CHO-sialyl Le(x) adhered efficiently to E-selectin, while adhesion of CHO-sulfo Le(x) was very poor despite the fact that near equal number of the ligands had been attached to the cell surface. In contrast, CHO-sulfo Le(x) adhered efficiently to P-selectin, while CHO-sialyl Le(x) adhered modestly to P-selectin. These results demonstrate that sialyl Le(x) and sulfo Le(x) structures on the cell surface differ substantially in their ability to adhere to E- and P-selectin.

Animals↗

Le(a) and Le(b) tissue glycosphingolipids.

BACKGROUND: The Le(a) and Le(b) antigens are synthesized by cells making exocrine secretions, but it is not well-defined where plasma and red cell Le(a) and Le(b) antigens are made. STUDY DESIGN AND METHODS: This study uses chromatographic and immunochemical methods to extract Le(a) and Le(b) glycolipids from human tissues. RESULTS: Le(a) and Le(b) antigens may be synthesized by both epithelial and mesodermal tissues. Expression of Lewis transferase activity was demonstrated in various tissues, confirming the epistatic interaction by Le and H genes to form the Leb antigen. CONCLUSION: The Le(a) and Le(b) substances found in plasma might originate from the epithelial cells of the digestive tract.

Epithelium↗

Neutrophil oxidative burst activation and the pattern of respiratory physiologic abnormalities in the fulminant post-traumatic adult respiratory distress syndrome.

The role of neutrophil oxidative burst activation (OBA) in the development of fulminant post-trauma adult respiratory distress syndrome (ARDS) was studied in 30 patients. Neutrophil (PMN) chemiluminescence (LE) was used as the index of OBA. Serially, for 8 days post-trauma, patient neutrophils (Pc) were studied in their own serum (Ps) normal serum (Ns), or Gey's solution (G). Ps was checked against normal neutrophils (Nc) for inhibition. LE was initiated by the addition of preopsonized zymosan to 1 x 10(6) PMN, the LE response monitored by luminometer, and the peak of the integral of LE recorded. Seven developed ARDS within the first 4 days; 12 patients developed sepsis (TS) but no ARDS, and 11 patients had uncomplicated trauma (TR). All ARDS showed increased LE (P less than 0.0001), at 48-96 hr. Patients without ARDS showed no significant increase in LE, although their mean injury severity (ISS) was the same. The ARDS LE response was mediated by activation of Pc [74%] with only a small but significant additional effect (6%) by ARDS serum (Ps): LE = 0.672 (Pc) + 0.24 [ARDS(Ps)] + 1343; N = 146, r2 0.733, P less than 0.0001. However, sera (Ps or Ns) was required, as incubation in G inhibited LE; [cells + s] greater than [cells + G], P less than 0.0001. LE is a biologic marker of ARDS, and the delay between injury and the LE indicated that initiation of ARDS may have therapeutic importance. Neutrophil activation in ARDS requires sera, but the ARDS effect appears mainly due to cells with only a small ARDS-specific serum-mediated role. The physiologic response to ARDS was evaluated by serial 8-hr studies of blood gases and pH; the respiratory index (RI) to pulmonary shunt (QS/QT) relationship, compliance (COMPL), and net fluid balance (DFLUID) PMN and platelet (PLAT) counts were also measured. Compared with TR and TS, the ARDS patients at 48-96 hr, showed increased RI, QS/QT, and DFluid requiring increased FiO2 and PEEP as COMPL and PLAT fell and LE rose. These changes were all simultaneously significant (P less than 0.05 to P less than 0.0001) by Bonferroni t-statistic applied to ANOVA. The clinical importance of these physiologic and biochemical responses was emphasized by the significantly (P less than 0.005) increased mortality in the ARDS patients. These data suggest that PMN LE and simple measures of respiratory function are early biologic markers of the development of fulminant post-traumatic ARDS and can be used to predict ARDS severity.

Adult↗

[Nucleophagocytosis in monocytes--an immunocytologic phenomenon of anti-dna-antibodies].

In SLE-sera antinuclear antibodies pervent normal nuclear degeneration, i.e. lysis of free nuclei. In sera with high titers, small rigid LE-bodies develop rapidly, which are soon phagocytized in toto by neutrophil granulocytes and monocytes. Classic LE-cells as well as Tart-cells are formed. The foreign appearance of the phagosomes in the monocytes is a result of the different digestive processes occuring in these phagocytes. With decreasing serum activity, the LE-bodies expand somewhat more and become viscous. From these nuclei, relatively small portions are phagocytized fairly late. Sjögren-cells of the granulocytic as well as the monocytic type are generated. Nucleophagocytosis only occurs in the presence of antinuclear antibodies, regardless of whether it leads to LE-cell, Tart-cell or partial phagocytosis, i.e. Sjögren-cells of the granulocytic or monocytic type. A non-specific nucleophagocytosis does not exist.

Antibodies, Anti-Idiotypic↗

Biological equivalence of natural bovine and recombinant human alpha-endothelial cell growth factors.

The cDNA encoding human alpha-endothelial cell growth factor (alpha-ECGF) has been engineered for high-level expression in Escherichia coli. Induction of bacterial cultures harboring the recombinant plasmid pMJ26 results in the appearance of a prominent 16-kDa polypeptide. This protein has been purified from bacterial lysates using a rapid, 2-step procedure employing heparin-Sepharose affinity based chromatography and reversed-phase high pressure liquid chromatography. Recombinant human alpha-ECGF was compared to bovine brain-derived alpha-ECGF in three biological assays: receptor binding on murine lung capillary endothelial cells (LE-II cells), stimulation of [3H]thymidine incorporation in LE-II cells, and stimulation of human umbilical vein endothelial cell proliferation. The results demonstrate that the recombinant human mitogen has the same biological potency as the bovine brain-derived material. Fluorescence spectroscopy was used to study the interaction between recombinant ECGF and heparin. Heparin-binding resulted in a 40% reduction in the intrinsic fluorescence of ECGF, consistent with a heparin-induced conformational change. The intrinsic fluorescence of ECGF also varied as a function of pH.

Amino Acids↗

Identification of the epidermal "Stiftchenzellen" of frog tadpoles by electron microscopy.

Differentiated surface epidermal cells observed in the skin of tadpoles of Rana temporaria by electron microscopy have been identified with the Stiftchenzellen originally described by Kölliker in 1885. The cells have apical microvilli or a single apical projection and appear to have synaptic associations with nerve fibres in the epidermis. The distribution, dimensions and structure of the cells are in agreement with descriptions from le cells are sensory in nature. In addition, there are fine structural resemblances to the gustatory cells of fish and of amphibians which suggest that the Stiftchenzellen are chemoreceptors.

Animals↗

The precipitin reaction between DNA and a serum factor in systemic lupus erythematosus.

The sera of certain patients with systemic lupus erythematosus contain an antibody-like substance capable of reacting with highly purified DNA preparations from widely divergent sources. Precipitin reactions have been demonstrated by double diffusion in agar and quantitative precipitin curves have been obtained. Complement was observed to be fixed in the reaction. Evidence was obtained that the serum factor possessed antigenic properties similar to those of gamma-globulins and migrated with this fraction on zone electrophoresis. The interaction of this factor with DNA exhibited certain specific characteristics which differ considerably from non-specific reactions between DNA and proteins in general. The DNA-precipitating factor appeared to be one of a number of related factors reacting with nuclear constituents of many different cells. It differed in certain respects from the "LE factor" which is responsible for the formation of "LE cells." The accumulated evidence, although not yet conclusive, favors the concept that the precipitating factor represents an antibody to DNA, and that it is one of a number of autoantibodies elicited in this disease.

Antibodies↗

[Urticarial vasculitis syndrome. A case report and review of the literature].

Urticarial vasculitis (UV) is a primary syndrome or a cutaneous vasculitic lesion occurring in the course of a collagen disease, as the systemic lupus erythematosus (LE). UV is a recently recognized disorder which affects most exclusively the female sex and may be differentiated from common (nonvasculitic) urticaria because it is characterized by inflammation and necrosis of blood vessels (vasculitis). UV and common urticaria may be induced by a variety of factors and pathogenetic mechanisms. It seems that a continuum exists, ranging from benign cutaneous lesions of urticaria to vasculitis with strong immunological involvement. On the basis of the clinical evaluation, two major groups of UV have been classified, the normocomplementemic, with a less severe clinical course, and the hypocomplementemic UV, a rare immune complex-mediated disorder related to LE as for as the similar pathogenesis and systemic involvement are concerned. A case of a young female patient with a unique syndrome characterized by crises of urticarial recurrent painful lesions of unknown origin, associated with angioedema of the tongue and soft palate, severe malaise, arthralgias and abdominal pain is reported. Dyspnea and cough were sometimes present, but fever was absent. Symptoms were caused by physical stress, heat or pressure and were little responsive to corticosteroids. Although skin biopsy did not prove with certainty the occurrence of vasculitis, clinical data and laboratory findings (normal levels of complement fractions, raised erythrocyte sedimentation rate, presence of cryoglobulins and ASMA, nDNA and AMA autoantibodies and absence of ANA autoantibodies and LE cells) suggest a normocomplementemic UV, excluding a classic LE. The family doctors and the internist must become familiar with the "lupus-like" syndromes, which include UV. In fact, a correct diagnosis of this syndrome is important because, although prognosis of UV may not be severe, the possibility exists of a systemic involvement (mainly renal) with progression to LE.

Adult↗

Lens epithelial cell mRNA. III. Elevated expression of macrophage migration inhibitory factor mRNA in galactose cataracts.

A lens epithelial (LE) cell cDNA clone, designated Clone 156, was isolated from a mature rat LE cDNA library by methods of subtractive hybridization. The cDNA sequence of Clone 156 was 521 nucleotides in length, excluding the poly(T)-tail, and it encoded an open reading frame of 115 amino acids. The translated protein shared extensive sequence similarities with macrophage migration inhibitory factor (MIF) from mouse lens and human T-cell lymphocytes. Northern blot hybridization showed that rat lens MIF mRNA is about 500 nucleotides in length and that its expression in mature rat lens is relatively low in comparison with that in other rat tissues. The expression of MIF mRNA in LE of normal rats and of rats treated by feeding a diet of 50% (w/w) galactose was studied by quantitative RT-PCR. The results showed that the expression of MIF mRNA in a 20-day galactosemic rat LE increased twelvefold as compared to that found in control LE. From the results of this study and from what we know about the locale of epithelial cell differentiation in mature rat lenses, it is being proposed that the increase in abundance of MIF mRNA in the cataractous rat lens is correlated with the enhanced proliferation of the undifferentiated epithelial cells.

Animals↗

[Thrombophlebitis of internal cerebral veins in a case of systemic lupus erythematosus (author's transl)].

A 28-year-old man suffered from recurrent facial exanthema, arthritis and stomatitis for ten years and died six months after a catatonic episode with terminal cerebral convulsions. Three years before his death high KBR-Antititers to Herpes simplex- and cytomegalic virus were observed, while Lupus-Erythematosus-Tests (LE-Tests) only became positive in the last months. At autopsy, changes compatible with Systemic Lupus Erythematosus (SLE) were found in the mitral valves, the spleen, and the kidneys. The brain displayed hemorrhagic infarction of the striate bodies and thrombophlebitis of the internal cerebral veins, the wall of which exhibited circumscribed infiltrations with numerous hematoxilin bodies and LE cells. This seems to be the first observation of LE-specific changes in the brain. The importance of cerebral vein affection in SLE involving the nervous system is stressed and a hypothesis submitted proposing the viral etiology of SLE.

Adult↗

[Jaccoud's type arthritis in a patient with unclassified connective tissue disease complicated with alveolar cell carcinoma].

The association between Jaccoud's type arthritis and systemic lupus erythematosus (SLE) is well recognized. Myositis, various arthropathy, and an assortment of miscellaneous connective tissue disorders have been described in association with malignancies. To date however Jaccoud's type arthritis has not been reported in unclassified connective tissue disease (UCTD) complicated with alveolar cell carcinoma (ACC). We describe a UCTD patient who presented LE cells, fluorescent antinuclear antibody (FANA), developing nonerosive, deforming arthritis and ACC. The case is a 59-year-old female who was admitted to our department in November, 1991 for exacerbated exertional dyspnea. She presented with UCTD of six years duration, characterized during follow-up by no history of rheumatic fever or Sjögren's syndrome, nor SLE. Five years later from onset of UCTD, her fingers developed a marked ulnar deviation, as well as pronounced swan-neck deformities. However, radiology did not show the marginal erosions of rheumatoid arthritis. A chest radiograph revealed a reticulogranular shadow in allover lung field, and a lung scintigram showed perfusion dominant mismatched defect in bilateral lower lung field. A transbronchial lung biopsy of B8a was diagnostic for ACC.

Adenocarcinoma, Bronchiolo-Alveolar↗

Systemic lupus erythematosus presenting as pleural effusion: report of a case.

Systemic lupus erythematosus (SLE) presenting as a pleural effusion in a young male is not common. This paper describes a 20-year-old man who was admitted to hospital with a spiking fever, chills and cough. A chest x-ray showed alveolar infiltration and a moderate right-sided pleural effusion. The patient was treated for parapneumonic effusion. Thoracentesis was performed and cytology of the aspirated fluid was initially interpreted as showing only numerous polymorphonuclear (PMN) leukocytes. However, in spite of antibiotic treatment the symptoms persisted. A careful review of the cytology specimen showed classic lupus erythematosus (LE) cells in addition to PMN cells. Subsequent investigation, including antinuclear antibodies titer, confirmed the diagnosis of LE pleurisy. Therapy with antibiotics was discontinued and treatment with prednisolone 20 mg daily was begun. There was a rapid clinical response including resolution of the fever and pleural effusion.

Adult↗

The base specificities of tomato ribonuclease (RNase LE) and its Asp44 mutant enzyme expressed from yeast cells.

RNase LE from cultured tomato cells is a member of the RNase T2 family. It is, however, distinguishable from RNase Rh from Rhizopus niveus, a typical RNase of the RNase T2 family, by its CD spectrum in the 200-250 nm region. In order to reinvestigate the base specificity of RNase LE and to study the role of Asn44 in RNase LE, which is considered to correspond to the base recognition site Asp51 of RNase Rh, RNase LE, and its Asp mutant at the 44th position were expressed from yeast cells with the same expression system as RNase Rh [K. Ohgi, et al., J. Biochem., 109, 776-785 (1991)]. RNase LE with four extra amino acid residues at the 2nd amino acid residue of mature RNase LE and its Asp44 mutant were secreted from yeast cells to give a yield of 10 mg/liter and 0.5 mg/liter culture broth, respectively. The expressed RNase LE (RNase RNAP LE) had the same characteristics as native RNase LE in the CD spectrum and specific activity. This is the first example of the expression of plant RNase from microbes and in sufficient amount to perform further enzymological research. The base specificity of RNase LE was guanylic acid preferential and that of N44D was changed to a more adenylic acid preference as compared to that of RNase LE. These experiments showed that Asn44 of RNase LE is crucial for base recognition as the case of Asp51 in RNASE Rh, and also suggested that the base recognition mechanism of RNase LE is very similar to that of RNase Rh.

Amino Acid Sequence↗

[Thymoma associated with myasthenia, erythroblastopenia, myositis and giant cell myocarditis. One case (author's transl)].

The authors report on a 76-year-old woman presenting with malignant thymoma associated with myasthenia, erythroblastopenia, myositis and giant cell myocarditis. Blood examination showed antinuclear antibodies, anti-platelet antibodies and LE cells. Only one similar case and 22 cases of thymoma associated with giant cell myocarditis have been published. It seems probable that thymomas plays a part in the pathogenesis of giant cell myocarditis.

Aged↗

Inductive capacity of living eye tissues from adult frogs.

The aim of the present work has been to demonstrate the inducing capacity of living homogenous undamaged tissues from adult frogs. Tissues from the eyes of adult frogs Xenopus laevis: retina (R), pigmented epithelium (PE), lens epithelium (LE), and the forebrain (B) as a control, were tested for their inducing capacity using early gastrula ectoderm. To exclude the possibility of ad-mixture of inducing cells in early gastrula ectoderm the transfilter induction technique was used throughout. The results show that the tissues used in the cases of most intense induction tended to induce similar cell types: both R and PE induce R + PE together with adjoining neural cells and secondary lens cells. LE induces lentoids (L) and B cells induce neuroids. In each case epidermis surrounds the explants filled with ectomesenchyme (EM) and melanophores (M). Immunofluorescence reactions clarified the nature of lens cells. This discovery indicates that cells of adult tissues continue throughout life producing substances which are able to promote the appearance of cells of the same type during development. Probably they also serve to mediate interrelations between cells of these tissues, regulating the stability of their differentiation in the adult state, as well.

Age Factors↗

Long-term preservation of canine bone marrow: in vitro studies.

In vitro studies were performed on canine bone marrow frozen with DMSO and stored in liquid nitrogen for 2 to 6 months. The results are compared with previously reported parallel in vivo experiments that demonstrated no loss of stem cells. When studies were performed immediately after thawing, there was no substantial drop in the count of nucleated cells and, except for megakaryocytes, there was no alteration of the bone marrow morphology. After two washes, and removal of DMSO, the nucleated cell count dropped to 50% of its previous value. Optic and electron microscopy showed severe damage in mature myeloid elements. In some instances, the cells had a condensed nucleus similar to the red-purple inclusion body of LE cells (as observed in systemic lupus erythematosus), and electron microscopy showed heavy chromatin clumping. On the other hand, both optic and electron microscopy showed a good preservation of lymphocytes, plasmocytes, and erythroid precursors. Two-hour DNA synthesis slightly dropped after storage, and this drop appeared more consistent when related to a constant volume of bone marrow (50 microliters) rather than to a constant number of nucleated cells (10(6)). In five instances frozen and thawed bone marrow was grown in short-term cultures, and analysis of 98 metaphases showed no major aberrations of the chromosomes and only 2% of minor aberrations, such as breakages and fragments. These data, compared with the results of previous in vivo experiments that showed no loss of stem cells after 5 months storage, suggest that stem cells are less sensitive to freezing and thawing injury than myeloid elements and/or that it might be safer for the thawed bone marrow not to be manipulated before infusion.

Animals↗