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Sample records for antineutrophil cytoplasm antibodies

  1. Antineutrophil cytoplasmic antibodies

    Directory of Open Access Journals (Sweden)

    G.D. Sebastiani

    2011-06-01

    Full Text Available Antineutrophil cytoplasmic antibodies (ANCA are predominantly IgG autoantibodies directed against constituents of primary granules of neutrophils and monocytes’ lysosomes. Although several antigenic targets have been identified, those ANCA directed to proteinase 3 or myeloperoxidase are clinically relevant, whereas the importance of other ANCA remains unknown. Both are strongly associated with small vessel vasculitides, the ANCA-associated vasculitides, which include Wegener’s granulomatosis, microscopic polyangiitis, and Churg-Strauss syndrome, and the localised forms of these diseases (eg, pauci-immune necrotising and crescentic glomerulonephritis. ANCA is a useful serological test to assist in diagnosis of small-vessel vasculitides. 85-95% of patients with Wegener’s granulomatosis, microscopic polyangiitis, and pauci-immune necrotising and crescentic glomerulonephritis have serum ANCA. ANCA directed to either proteinase 3 or myeloperoxidase are clinically relevant, yet the relevance of other ANCA remains unknown. Besides their diagnostic potential, ANCA might be valuable in disease monitoring. In addition, data seem to confirm the long-disputed pathogenic role of these antibodies. There is increasing evidence that myeloperoxidase- ANCA are directly involved in the pathogenesis of necrotizing vasculitis. This is less clear for proteinase 3-ANCA, markers for Wegener’s granulomatosis. With respect to proteinase 3-ANCA, complementary proteinase 3, a peptide translated from the antisense DNA strand of proteinase 3 and homologous to several microbial peptides, may be involved in induction of proteinase 3-antineutrophil cytoplasmic autoantibodies.

  2. Necrotizing and Crescentic Lupus Nephritis with Antineutrophil Cytoplasmic Antibody Seropositivity

    OpenAIRE

    Nasr, Samih H.; D'Agati, Vivette D; Park, Hye-Ran; Sterman, Paul L.; Goyzueta, Juan D.; Dressler, Robert M.; Hazlett, Shawn M.; Pursell, Robert N.; Caputo, Christopher; Markowitz, Glen S.

    2008-01-01

    Background and objectives: Lupus nephritis is a classic immune complex glomerulonephritis. In contrast, antineutrophil cytoplasmic antibodies are associated with necrotizing and crescentic glomerulonephritis, in the absence of significant immune deposits. Antineutrophil cytoplasmic antibodies are detected by indirect immunofluorescence in 20% of patients with systemic lupus erythematosus. We report 10 cases of necrotizing and crescentic lupus nephritis with antineutrophil cytoplasmic antibody...

  3. Clinical analyses of perinuclear antineutrophil cytoplasmic antibody associated hypertrophic pachymeningitis

    Institute of Scientific and Technical Information of China (English)

    赵鹏

    2013-01-01

    Objective To explore the clinical characteristics,diagnosis,treatment and prognosis of perinuclear antineutrophil cytoplasmic antibody(p-ANCA)associated hypertrophic pachymeningitis.Methods A retrospective study

  4. Animal models of antineutrophil cytoplasm antibody-associated vasculitis.

    LENUS (Irish Health Repository)

    Salama, Alan D

    2012-01-01

    To provide an update on the experimental models that have been developed recapitulating clinical antineutrophil cytoplasm antibody (ANCA) associated vasculitis. The application of the models in the study of pathogenesis, and the therapeutic implications of this, are covered in the article by van Timmeren and Heeringa in this issue.

  5. Anti-neutrophil cytoplasmic antibodies in idiopathic inflammatory disorders

    NARCIS (Netherlands)

    Mulder, Alide Heleen Leontine

    1994-01-01

    This thesis studies the prevalence of anti-neutrophil cytoplasmic antibodies (ANCA) in idiopathic inflammatory disorders in which GS-ANA were described during the 60s: rheumatoid arthritis, autoimmune liver diseases, inflammatory bowel disease, and juvenile chronic arthritis. In all of these disorde

  6. Evaluation of antineutrophil cytoplasmic antibody seroconversion induced by minocycline, sulfasalazine, or penicillamine

    NARCIS (Netherlands)

    Choi, HK; Slot, MC; Pan, GL; Weissbach, CA; Niles, JL; Merkel, PA

    2000-01-01

    Objective, Case reports have suggested that minocycline, sulfasalazine, and penicillamine are associated with antineutrophil cytoplasmic antibody (ANCA)-positive vasculitis, This study evaluated ANCA seroconversion due to these agents in serum samples prospectively collected in randomized, double-bl

  7. Anti-neutrophil cytoplasmic antibodies in rheumatoid arthritis: two case reports and review of literature

    Directory of Open Access Journals (Sweden)

    Spoerl David

    2012-12-01

    Full Text Available Abstract Background Anti-neutrophil cytoplasmic antibodies are typically detected in anti-neutrophil cytoplasmic antibody associated vasculitis, but are also present in a number of chronic inflammatory non-vasculitic conditions like rheumatoid arthritis. Rare cases of granulomatosis with polyangiitis (formerly known as Wegener’s granulomatosis, a vasculitic disorder frequently associated with the presence of anti-neutrophil cytoplasmic antibodies in patients with rheumatoid arthritis have been described in literature. Case presentation We report two middle-aged female patients with rheumatoid arthritis who developed anti-neutrophil cytoplasmic antibodies and symptoms reminiscent of granulomatosis with polyangiitis. Despite the lack of antibodies specific for proteinase 3 and the absence of a classical histology, we report a probable case of granulomatosis with polyangiitis in the first patient, and consider rheumatoid vasculitis in the second patient. Conclusion Taken together with previous reports, these cases highlight that anti-neutrophil cytoplasmic antibodies have to be evaluated very carefully in patients with rheumatoid arthritis. In this context, anti-neutrophil cytoplasmic antibodies detected by indirect immunofluorescence appear to have a low diagnostic value for granulomatosis with polyangiitis. Instead they may have prognostic value for assessing the course of rheumatoid arthritis.

  8. Anti-neutrophil cytoplasmic antibodies stimulate release of neutrophil microparticles.

    LENUS (Irish Health Repository)

    Hong, Ying

    2012-01-01

    The mechanisms by which anti-neutrophil cytoplasmic antibodies (ANCAs) may contribute to the pathogenesis of ANCA-associated vasculitis are not well understood. In this study, both polyclonal ANCAs isolated from patients and chimeric proteinase 3-ANCA induced the release of neutrophil microparticles from primed neutrophils. These microparticles expressed a variety of markers, including the ANCA autoantigens proteinase 3 and myeloperoxidase. They bound endothelial cells via a CD18-mediated mechanism and induced an increase in endothelial intercellular adhesion molecule-1 expression, production of endothelial reactive oxygen species, and release of endothelial IL-6 and IL-8. Removal of the neutrophil microparticles by filtration or inhibition of reactive oxygen species production with antioxidants abolished microparticle-mediated endothelial activation. In addition, these microparticles promoted the generation of thrombin. In vivo, we detected more neutrophil microparticles in the plasma of children with ANCA-associated vasculitis compared with that in healthy controls or those with inactive vasculitis. Taken together, these results support a role for neutrophil microparticles in the pathogenesis of ANCA-associated vasculitis, potentially providing a target for future therapeutics.

  9. ANTINEUTROPHIL CYTOPLASMIC ANTIBODIES - A STILL-GROWING CLASS OF AUTOANTIBODIES IN INFLAMMATORY DISORDERS

    NARCIS (Netherlands)

    KALLENBERG, CGM; MULDER, AHL; TERVAERT, JWC

    1992-01-01

    Antineutrophil cytoplasmic antibodies (ANCA) have been described as sensitive and specific markers for active Wegener's granulomatosis (WG). ANCA in WG produce a characteristic cytoplasmic staining pattern of neutrophils (c-ANCA) and are directed against proteinase 3 (Pr3), a serine protease from th

  10. Clinical Outcomes of Remission Induction Therapy for Severe Antineutrophil Cytoplasmic Antibody-Associated Vasculitis

    NARCIS (Netherlands)

    Miloslavsky, E. M.; Specks, U.; Merkel, P. A.; Seo, P.; Spiera, R.; Langford, C. A.; Hoffman, G. S.; Kallenberg, C. G. M.; St Clair, E. W.; Tchao, N. K.; Viviano, L.; Ding, L.; Sejismundo, L. P.; Mieras, K.; Ikle, D.; Jepson, B.; Mueller, M.; Brunetta, P.; Allen, N. B.; Fervenza, F. C.; Geetha, D.; Keogh, K.; Kissin, E. Y.; Monach, P. A.; Peikert, T.; Stegeman, C.; Ytterberg, S. R.; Stone, J. H.

    2013-01-01

    Objective. To evaluate the reasons that complete remission is not achieved or maintained with original treatment in some patients with antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) treated with rituximab (RTX) or with cyclophosphamide/azathioprine (CYC/AZA). Methods. The Rit

  11. Propylthiouracil induced anti-neutrophil cytoplasmic antibody-associated vasculitis with bone marrow plasmacytosis andgranulocytopenia

    Institute of Scientific and Technical Information of China (English)

    Abdullah Ozkok

    2009-01-01

    @@ Antithyroid drugs are molecules known as thionamides that inhibit thyroid hormone synthesis by interfering with thyroid peroxidase mediated iodination of tyrosine residues in thyroglobulin. These extensively used drugs are associated with a variety of well-known side effects such as anti-neutrophil cytoplasmic antibody (ANCA)-positive vasctilitis, granulocytopenia and aplastic anemia.

  12. Menopause and Primary Ovarian Insufficiency in Women Treated for Antineutrophil Cytoplasmic Antibody-Associated Vasculitides

    NARCIS (Netherlands)

    Tuin, Janneke; Sanders, Jan-Stephan F.; van Beek, Andre P.; Hoek, Annemieke; Stegeman, Coen A.

    2016-01-01

    ObjectiveOne of the side effects of cyclophosphamide is earlier menopause and primary ovarian insufficiency. This study was undertaken to investigate the onset of menopause and the incidence of primary ovarian insufficiency in women with antineutrophil cytoplasmic antibody-associated vasculitis (AAV

  13. Pediatric Inflammatory Bowel Disease with Cytoplasmic Staining of Antineutrophil Cytoplasmic Antibodies

    Directory of Open Access Journals (Sweden)

    Omar I. Saadah

    2013-01-01

    Full Text Available Background. It is unusual for the antineutrophil cytoplasmic antibody with cytoplasmic pattern (cANCA to present in patients with inflammatory bowel disease (IBD without vasculitis. The purpose of this study was to describe the occurrence and characteristics of pediatrics IBD with cANCA. Methods. A retrospective review of pediatric IBD associated with cANCA serology in patients from King Abdulaziz University Hospital, Saudi Arabia, between September 2002 and February 2012. Results. Out of 131 patients with IBD screened for cANCAs, cANCA was positive in 7 (5.3% patients of whom 4 had ulcerative colitis and 3 had Crohn's disease. The median age was 8.8 years (2–14.8 years. Six (86% were males. Of the 7 patients, 5 (71% were Saudi Arabians and 2 were of Indian ethnicity. The most common symptoms were diarrhea, abdominal pain, weight loss, and rectal bleeding. None had family history or clinical features suggestive of vasculitis involving renal and respiratory systems. No difference in the disease location or severity was observed between cANCA positive and cANCA negative patients apart from male preponderance in cANCA positive patients. Conclusion. The occurrence of cANCA in pediatric IBD is rare. Apart from male preponderance, there were no peculiar characteristics for the cANCA positive patients.

  14. Differentiation of Behcet's disease from inflammatory bowel diseases: Anti-saccharomyces cerevisiae antibody and anti-neutrophilic cytoplasmic antibody

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    The differential diagnosis of Behcet's disease (BD) from inflammatory bowel disease (IBD) is sometimes difficult and challenging. Hereby, we suggested the utility of anti-saccharomyces cerevisiae antibody (ASCA) and anti-neutrophilic cytoplasmic antibody (p-ANCA) in the differential diagnosis of BD from IBD.

  15. Pulse versus daily oral cyclophosphamide for induction of remission in antineutrophil cytoplasmic antibody-associated vasculitis: a randomized trial

    DEFF Research Database (Denmark)

    de Groot, Kirsten; Harper, Lorraine; Jayne, David R W;

    2009-01-01

    BACKGROUND: Current therapies for antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis are limited by toxicity. OBJECTIVE: To compare pulse cyclophosphamide with daily oral cyclophosphamide for induction of remission. DESIGN: Randomized, controlled trial. Random assignments were...

  16. Plasma exchange in antineutrophil cytoplasmic antibody-associated vasculitis--a 25-year perspective

    DEFF Research Database (Denmark)

    Szpirt, Wladimir M

    2015-01-01

    Demonstration of a pathogenic role for antineutrophil cytoplasmic antibodies (ANCA) underlies the scientific rationale for plasma exchange (PLEX) in the treatment of ANCA-associated vasculitis (AAV). Most clinical evidence of efficacy concerns the use of PLEX for the recovery of renal function in...... in severe nephritis, when used in conjunction with immunosuppressive drug therapy. The development of PLEX for this indication, the strength of the clinical trial evidence supporting its use, its roles in other AAV indications and ongoing research are discussed....

  17. Plasma exchange and glucocorticoid dosing in the treatment of anti-neutrophil cytoplasm antibody associated vasculitis (PEXIVAS)

    DEFF Research Database (Denmark)

    Walsh, Michael; Merkel, Peter A; Peh, Chen Au;

    2013-01-01

    Granulomatosis with polyangiitis (GPA, Wegener's) and microscopic polyangiitis (MPA) are small vessel vasculitides collectively referred to as anti-neutrophil cytoplasm antibody-associated vasculitis (AAV). AAV is associated with high rates of morbidity and mortality due to uncontrolled disease and...

  18. Brief Report : Menopause and Primary Ovarian Insufficiency in Women Treated for Antineutrophil Cytoplasmic Antibody-Associated Vasculitides

    NARCIS (Netherlands)

    Tuin, Janneke; Sanders, Jan-Stephan F; van Beek, André P; Hoek, Annemieke; Stegeman, Coen A

    2016-01-01

    OBJECTIVE: One of the side effects of cyclophosphamide is earlier menopause and primary ovarian insufficiency. This study was undertaken to investigate the onset of menopause and the incidence of primary ovarian insufficiency in women with antineutrophil cytoplasmic antibody-associated vasculitis (A

  19. Circulating microRNA expression pattern separates patients with anti-neutrophil cytoplasmic antibody-associated vasculitis from healthy controls

    DEFF Research Database (Denmark)

    Skoglund, C.; Carlsen, A.; Weiner, M.;

    2015-01-01

    Objective. Antineutrophil cytoplasmic antibody-associated vasculitis (AAV) has an unpredictable course and better biomarkers are needed. Micro-RNAs in body fluids are protected from degradation and might be used as biomarkers for diagnosis and prognosis, here we explore the potential in AAV...

  20. Unusual cause of flare in antineutrophil cytoplasmic antibody-associated vasculitis.

    Science.gov (United States)

    Glanville, James Robert William; Penn, Henry

    2016-01-01

    A 53-year-old woman attended for a routine outpatient appointment for follow-up of antineutrophil cytoplasmic antibody-positive vasculitis. Her disease had relapsed despite appropriate medical management with mycophenolate mofetil (MMF), as evidenced by rising acute phase response and antimyeloperoxidase titre with ongoing symptoms. On further questioning, she had been taking oral charcoal as part of a detoxification diet, which we postulate was causing significantly impaired MMF absorption. This case report summarises the presentation and highlights the importance of a thorough drug history, and should prompt the reader to keep an open mind with regard to drug interactions and treatment regimen adherence when treatment is, unexpectedly, seemingly failing. PMID:26746838

  1. Antineutrophil Cytoplasmic Antibody Induction due to Infection: A Patient with Infective Endocarditis and Chronic Hepatitis C.

    Science.gov (United States)

    Kamar, Fareed B; Hawkins, T Lee-Ann

    2016-01-01

    While antineutrophil cytoplasmic antibody (ANCA) is often used as a diagnostic marker for certain vasculitides, ANCA induction in the setting of infection is much less common. In the case of infective endocarditis, patients may present with multisystem disturbances resembling an autoimmune process, cases that may be rendered even trickier to diagnose in the face of a positive ANCA. Though not always straightforward, distinguishing an infective from an inflammatory process is pivotal in order to guide appropriate therapy. We describe an encounter with a 43-year-old male with chronically untreated hepatitis C virus infection who featured ANCA positivity while hospitalized with acute bacterial endocarditis. His case serves as a reminder of two of the few infections known to uncommonly generate ANCA positivity. We also summarize previously reported cases of ANCA positivity in the context of endocarditis and hepatitis C infections. PMID:27366166

  2. The clinical presentation and therapy of diseases related to anti-neutrophil cytoplasmic antibodies (ANCA).

    Science.gov (United States)

    Weiner, Maria; Segelmark, Mårten

    2016-10-01

    Anti-neutrophil cytoplasmic antibodies (ANCA) are a family of autoantibodies that react with proteins predominantly expressed in cytoplasmic granules of polymorphonuclear neutrophil granulocytes (PMNs). ANCA was initially detected using indirect immunofluorescence, allowing for different patterns such as p-ANCA (perinuclear) and c-ANCA (cytoplasmic) to be distinguished. Today it is common to detect the antibodies by immunochemical assays such as ELISA using purified proteins as antigens. The strongest association with ANCA is found in the pauci-immune small vessel vasculitides granulomatosis with polyangiitis (GPA) and microscopic polyangiitis (MPA). There is compelling evidence that ANCA contributes to the pathogenesis in these conditions. ANCA also occurs in 30%-40% of patients with eosinophilic granulomatosis with polyangiitis (EGPA) and anti-GBM disease, but is uncommon in other forms of vasculitis. ANCA with different specificities have been described with varying frequencies in diseases such as systemic lupus erythematosus (SLE), rheumatoid arthritis, inflammatory bowel disease, endocarditis, chronic infections and hematopoietic malignancies. ANCA can also develop as an adverse event during pharmacological treatment. These entities are treated quite differently, with therapies ranging from immunosuppressive agents over antibiotics to simply removing the causative drug. A positive ANCA test thus requires a careful diagnostic work-up.

  3. The clinical presentation and therapy of diseases related to anti-neutrophil cytoplasmic antibodies (ANCA).

    Science.gov (United States)

    Weiner, Maria; Segelmark, Mårten

    2016-10-01

    Anti-neutrophil cytoplasmic antibodies (ANCA) are a family of autoantibodies that react with proteins predominantly expressed in cytoplasmic granules of polymorphonuclear neutrophil granulocytes (PMNs). ANCA was initially detected using indirect immunofluorescence, allowing for different patterns such as p-ANCA (perinuclear) and c-ANCA (cytoplasmic) to be distinguished. Today it is common to detect the antibodies by immunochemical assays such as ELISA using purified proteins as antigens. The strongest association with ANCA is found in the pauci-immune small vessel vasculitides granulomatosis with polyangiitis (GPA) and microscopic polyangiitis (MPA). There is compelling evidence that ANCA contributes to the pathogenesis in these conditions. ANCA also occurs in 30%-40% of patients with eosinophilic granulomatosis with polyangiitis (EGPA) and anti-GBM disease, but is uncommon in other forms of vasculitis. ANCA with different specificities have been described with varying frequencies in diseases such as systemic lupus erythematosus (SLE), rheumatoid arthritis, inflammatory bowel disease, endocarditis, chronic infections and hematopoietic malignancies. ANCA can also develop as an adverse event during pharmacological treatment. These entities are treated quite differently, with therapies ranging from immunosuppressive agents over antibiotics to simply removing the causative drug. A positive ANCA test thus requires a careful diagnostic work-up. PMID:27481040

  4. Risk factors for relapse of antineutrophil cytoplasmic antibody-associated vasculitis

    NARCIS (Netherlands)

    Walsh, Michael; Flossmann, Oliver; Berden, Annelies; Westman, Kerstin; Hoglund, Peter; Stegeman, Coen; Jayne, David

    2012-01-01

    Objective To determine the association between characteristics at diagnosis and the time to first relapse in a large cohort of patients with antineutrophil cytoplasmic antibodyassociated vasculitis (AAV). Methods. We studied long-term followup data from 4 clinical trials that included newly diagnose

  5. A rare case of recurrent pregnancy loss associated with high-titer positivity for perinuclear anti-neutrophilic cytoplasmic antibodies

    Directory of Open Access Journals (Sweden)

    Akhila Vasudeva

    2012-01-01

    Full Text Available We present a case of recurrent pregnancy loss associated with unusual constellation of utoimmunity-related features such as hypertension, severe hrombocytopenia, hypothyroidism and persistent high titers of perinuclear antineutrophilic cytoplasmic antibodies. Her clinical features did not fit into a particular diagnosis of vasculitides, systemic lupus erythematosis (SLE or other known autoimmune diseases where this autoantibody is found in high titers. We report the unusual association of this autoantibody with recurrent early fetal demise in this case.

  6. Sustained Remission of Antineutrophil Cytoplasmic Antibody-Mediated Glomerulonephritis and Nephrotic Syndrome in Mixed Connective Tissue Disease

    OpenAIRE

    Konstantinov, Konstantin N.; Harris, Alexis A.; Barry, Marc; Murata, Glen H; Tzamaloukas, Antonios H.

    2013-01-01

    A woman diagnosed with mixed connective tissue disease (MCTD) developed an anti-myeloperoxidase (MPO) antineutrophil cytoplasmic antibody (ANCA) and nephrotic syndrome with normal serum creatinine. Percutaneous kidney biopsy showed pauci-immune glomerulonephritis with superimposed immune complex deposition. After treatment with cyclophophamide and prednisone, proteinuria decreased progressively to a level of 0.4 g/g creatinine, ANCA became undetectable, while serum creatinine remained normal ...

  7. A subset of ulcerative colitis with positive proteinase-3antineutrophil cytoplasmic antibody

    Institute of Scientific and Technical Information of China (English)

    Jin Xu; Chuan-Hua Yang; Xiao-Yu Chen; Xu-Hang Li; Min Dai; Shu-Dong Xiao

    2008-01-01

    A small subset of patients with active ulcerative colitis is non-responsive to major known non-biological therapies.We reported 5 patients with positive serum proteinase-3 antineutrophil cytoplasmic antibody (PR3-ANCA) and tried to (1) identify the common clinical features of these patients; (2) investigate the efficacy of a novel therapy using a Chinese medicine compound; and (3) attract more gastroenterologists to be engaged in further study of this subset of patients. The common manifestations of disease in these 5 patients included recurrent bloody diarrhea and inflammatory lesions involving the entire colorectal mucosa. Initial treatment with intravenous methylprednisolone successfully induced remission.Four of these 5 patients were steroid-dependence,and immunosuppressants, such as azathioprine and cyclophosphamide, were ineffective. In 3 patients,only the particular Chinese medicine compound could induce and maintain remission. One patient underwent colectomy. No vascular inflammatory lesions were found by histopathological examination. Although more cases are needed for confirmation, our study indicates that ulcerative colitis with positive PR3-ANCA may belong to a subtype of refractory ulcerative colitis. The particular Chinese medicine compound used in our study is by far the most effective in the management of these patients,with additional advantages of having no noticeable sideeffects and less financial burden.

  8. CpG oligodeoxynucleotide stimulates production of anti-neutrophil cytoplasmic antibodies in ANCA associated vasculitis

    Directory of Open Access Journals (Sweden)

    Sarvestani Ghafar

    2008-07-01

    Full Text Available Abstract Background Wegener's Granulomatosis and Microscopic Polyangiitis are life-threatening systemic necrotizing vasculitides of unknown aetiology. The appearance of circulating antibodies to neutrophil cytoplasmic antigens (ANCA is strongly associated with the development of the disease. A link between infection and disease has long been suspected, and the appearance of ANCA antibodies has been reported following bacterial and viral infections. The depletion of circulating B cells with monoclonal antibody therapy can induce remission, and this observation suggests a pathogenic role for B cells in this disease. As bacterial DNA is known to induce B cell proliferation and antibody production via TLR-9 stimulation, we have explored the possibility that unmethylated CpG oligodeoxynucleotide, as found in bacterial and viral DNA, may play a role in stimulating circulating autoreactive B cells to produce ANCA in patients with vasculitis. Results We have confirmed that unmethylated CpG oligonucleotide is a potent stimulator of antibody production by PBMC in vitro. The stimulation of PBMC with CpG oligonucleutides resulted in the production of similar amounts of IgG in both ANCA+ patients and normal controls. In spite of this, PR3 ANCA+ patients synthesised significantly higher amount of IgG ANCA than normal controls. In MPO ANCA+ patients, there was a tendency for patients to produce higher amount of ANCA than controls, however, the difference did not reach significance. Furthermore, we were able to detect circulating MPO-reactive B cells by ELISpot assay from the peripheral blood of 2 MPO+ ANCA vasculitis patients. Together, this indicates that circulating anti-neutrophil autoreactive B cells are present in ANCA+ vasculitis patients, and they are capable of producing antibodies in response to CpG stimulation. Of note, CpG also induced the production of the relevant autoantibodies in patients with other types of autoimmune diseases. Conclusion

  9. Anti-neutrophil cytoplasm autoantibodies (ANCA) in autoimmune liver diseases

    NARCIS (Netherlands)

    Roozendaal, C.; Kallenberg, Cees

    1999-01-01

    Anti-neutrophil cytoplasm antibodies (ANCA) are autoantibodies directed against cytoplasmic constituents of neutrophil granulocytes and monocytes. ANCA have been detected in serum from patients with inflammatory bowel diseases (mainly ulcerative colitis) and autoimmune mediated liver diseases (mainl

  10. Antineutrophil cytoplasmic antibodies in bile are associated with disease activity in primary sclerosing cholangitis.

    Science.gov (United States)

    Lenzen, Henrike; Weismüller, Tobias J; Negm, Ahmed A; Wlecke, Jenny; Loges, Stephanie; Strassburg, Christian P; Manns, Michael P; Lankisch, Tim O

    2013-10-01

    OBJECTIVE. Primary sclerosing cholangitis (PSC) is an autoimmune cholestatic liver disease of unknown etiology. The role of antineutrophil cytoplasmic antibodies (ANCAs) in the serum of patients with PSC remains unclear. We hypothesized that ANCA may be detectable in bile, potentially providing diagnostic and prognostic information. METHODS. Serum and bile were prospectively collected during endoscopic retrograde cholangiography (ERC) in 72 patients with PSC and other non-PSC obstructive biliary diseases. ANCA measurements were performed by indirect immunofluorescence (IIF). RESULTS. Immunoglobulin G (IgG) ANCA was detected significantly more often in the bile of PSC patients (15/39; 38%) than without (2/33; 6%) (p = 0.001). IgG ANCA in bile was associated with a ten times higher risk of PSC (p = 0.005). In addition, IgG ANCA positivity in bile was associated with the presence of dominant strictures (p = 0.03), cholangiographic severity (p = 0.004), number of ERC (p = 0.01) and interventions performed (p = 0.03). However, IgG ANCA in bile did not correlate with transplantation, cholangiocarcinoma or death. No association was observed between ANCA positivity in sera and ANA and ASCA positivity in sera or bile with the above-mentioned clinical features. CONCLUSIONS. The presence of ANCA in the bile of patients with PSC is a novel finding and highly suggestive of PSC. Biliary IgG ANCA correlates with the severity of bile duct strictures and the ensuing number of ERCs and interventions. Therefore, a positive ANCA status in bile may serve as a diagnostic and prognostic marker of the disease progression and biliary complications.

  11. [Myeloperoxidase antineutrophil cytoplasmic antibody (MPO-ANCA) -associated glomerulonephritis with acute pancreatitis: a case report].

    Science.gov (United States)

    Iida, Takeshi; Amari, Yoshifumi; Yurugi, Takatomi; Nakajima, Fumitaka

    2015-01-01

    We report here a case of a 64-year-old woman with myeloperoxidase antineutrophil cytoplasmic antibody (MPO-ANCA) -associated glomerulonephritis who developed acute pancreatitis. The patient was admitted to our hospital because of abnormal urinalysis findings, edema, and progressive renal failure. Laboratory studies showed a high white blood cell count (11,570/μL), anemia (hemoglobin 7.8 g/dL), and elevated serum creatinine (2.36 mg/dL) and C-reactive protein (12.20 mg/dL) levels. Furthermore, the MPO-ANCA titer was very high (1,625 U/mL, normal range low back pain. Computed tomography (CT) revealed postbiopsy hemorrhage. Thereafter, the patient's symptoms and laboratory studies gradually worsened. A repeat CT performed a few days later revealed no changes in the perirenal hematoma; however, an enlarged pancreas head was incidentally observed. There was no obvious cause of acute pancreatitis, and MPO-ANCA-associated vasculitis, although rare, was suspected as the cause. We initiated prednisolone pulse therapy for vasculitis along with the administration of nafamostat mesilate and ulinastatin for acute pancreatitis. Subsequently, the levels of pancreatic enzymes gradually increased, but several days later, abdominal magnetic resonance imaging showed improvement in the pancreas head. The pancreatitis gradually resolved over time. Acute pancreatitis occurring concurrently with MPO-ANCA-associated glomerulonephritis is extremely rare. To our knowledge, only a few such cases have been reported and have suggested that steroid therapy may play a role in triggering pancreatic involvement. In our case, however, an enlarged pancreas head was observed before steroid therapy was initiated. Therefore, we consider our case to be very rare. PMID:26126336

  12. Anti-neutrophil cytoplasmic antibody-associated Pauci-immune crescentic glomerulonephritis complicating Sjögren's syndrome.

    Science.gov (United States)

    Wang, Wei-Jei; Wu, Hau-Shin; Chu, Tzong-Shinn

    2011-07-01

    Sjögren's syndrome is a chronic autoimmune disease, characterized by specific autoimmune antibodies anti-Ro and anti-La, and it can involve multiple organs, such as the kidneys, lungs, muscles, and nervous system. The most common renal complication of Sjögren's syndrome is tubulointerstitial nephritis, and glomerulonephritis is relatively uncommon. We report the case of an 86-year-old man presenting with recurrent fever, poor appetite, decreased salivary secretion, and body weight loss. Laboratory investigation revealed that serum creatinine was 4.2 mg/dL, proteinuria was 3+, and there was microscopic hematuria. Positive perinuclear anti-neutrophil cytoplasmic antibody, anti-Ro, and anti-La antibodies were detected. Renal biopsy showed crescentic glomerulonephritis with scanty immune complex deposition. The patient was diagnosed with primary Sjögren's syndrome complicated with rapidly progressive glomerulonephritis with positive anti-neutrophil cytoplasmic antibody. Unlike the patients of other case reports, our patient's renal function did not recover after immunosuppressant treatment, and he finally received long-term hemodialysis. Pauci-immune glomerulonephritis is a rare renal complication of Sjögren's syndrome, and progress to renal failure in such patients is possible.

  13. Spotlight on rituximab in the treatment of antineutrophil cytoplasmic antibody-associated vasculitis: current perspectives

    Directory of Open Access Journals (Sweden)

    Moog P

    2015-11-01

    Full Text Available Philipp Moog, Klaus Thuermel Abteilung für Nephrologie, Klinikum rechts der Isar, Technische Universität München, Munich, Germany Abstract: A 54-year-old patient presented to his general practitioner because of strong muscle pain in both thighs. Inflammatory parameters (CRP 16.3 mg/dL and white blood cells (15 g/L were elevated. The patient reported a weight loss of 10 kg in 4 weeks. There was no fever or any other specific symptoms. Urine dipstick examination and computed tomography of the chest were unremarkable. Because of increasing symptoms, the patient was referred to our department. Magnetic resonance tomography showed diffuse inflammatory changes of the muscles of both thighs. Neurological examination and electrophysiology revealed axonal sensorimotor neuropathy and ground-glass opacities of both lungs had occurred. Serum creatinine increased to 229 µmol/L within a few days, with proteinuria of 3.3 g/g creatinine. Kidney biopsy showed diffuse pauci-immune proliferative glomerulonephritis. Proteinase 3-specific antineutrophil cytoplasmic antibodies were markedly increased. Birmingham Vasculitis Activity Score was 35. Within 2 days, serum creatinine further increased to 495 µmol/L. Plasma exchange, high-dose glucocorticosteroids, and hemodialysis were started. The patient received cyclophosphamide 1 g twice and rituximab 375 mg/m2 four times according to the RITUXVAS protocol. Despite ongoing therapy, hemodialysis could not be withdrawn and had to be continued over 3 weeks until diuresis normalized. Glucocorticosteroids were tapered to 20 mg after 2 months, and serum creatinine was 133 µmol/L. However, nephritic urinary sediment reappeared. Another dose of 1 g cyclophosphamide was given, and glucocorticosteroids were raised for another 4 weeks. After 6 months, the daily prednisolone dose was able to be tapered to 5 mg. Serum creatinine was 124 µmol/L, proteinuria further decreased to 382 mg/g creatinine, and the Birmingham

  14. Combination therapy with rituximab and cyclophosphamide in the treatment of anti-neutrophil cytoplasmic antibodies (ANCA) positive pulmonary hemorrhage: case report

    OpenAIRE

    Lehman Thomas JA; Baird Emily M; Worgall Stefan

    2011-01-01

    Abstract Anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) with pulmonary hemorrhage is rare in childhood. Standard treatment includes corticosteroids and cyclophosphamide (CYC), which is associated with a high level of toxicity. We report a white female with ANCA positive pulmonary hemorrhage who was treated with cyclophosphamide (CYC) and rituximab (RTX) combination therapy.

  15. Systemic Lupus Erythematosus and Antineutrophil Cytoplasmic Antibody-Associated Vasculitis Overlap Syndrome in Patients With Biopsy-Proven Glomerulonephritis

    Science.gov (United States)

    Jarrot, Pierre-Andre; Chiche, Laurent; Hervier, Baptiste; Daniel, Laurent; Vuiblet, Vincent; Bardin, Nathalie; Bertin, Daniel; Terrier, Benjamin; Amoura, Zahir; Andrés, Emmanuel; Rondeau, Eric; Hamidou, Mohamed; Pennaforte, Jean-Loup; Halfon, Philippe; Daugas, Eric; Dussol, Bertrand; Puéchal, Xavier; Kaplanski, Gilles; Jourde-Chiche, Noemie

    2016-01-01

    Abstract The aim of the study was to report the clinical, biological, and pathological characteristics of patients with glomerulonephritis (GN) secondary to systemic lupus erythematosus (SLE)/antineutrophil cytoplasmic antibodies (ANCA)-associated vasculitis (AAV) overlap syndrome. A nationwide survey was conducted to identify cases of SLE/AAV overlap syndrome. Data were collected from SLE and AAV French research groups. Inclusion criteria were diagnosis of both SLE and AAV according to international classification criteria and biopsy-proven GN between 1995 and 2014. Additional cases were identified through a systematic literature review. A cohort of consecutive biopsy-proven GN was used to study the prevalence of overlapping antibodies and/or overlap syndrome. The national survey identified 8 cases of SLE/AAV overlap syndrome. All patients were female; median age was 40 years. AAV occurred before SLE (n = 3), after (n = 3), or concomitantly (n = 2). Six patients had rapidly progressive GN and 3/8 had alveolar hemorrhage. All patients had antinuclear antibodies (ANA); 7/8 had p-ANCA antimyeloperoxidase (MPO) antibodies. Renal biopsies showed lupus nephritis (LN) or pauci-immune GN. Remission was obtained in 4/8 patients. A literature review identified 31 additional cases with a similarly severe presentation. In the GN cohort, ANCA positivity was found in 30% of LN, ANA positivity in 52% of pauci-immune GN, with no correlation with pathological findings. The estimated prevalence for SLE/AAV overlap syndrome was 2/101 (2%). In patients with GN, SLE/AAV overlap syndrome may occur but with a low prevalence. Most patients have an aggressive renal presentation, with usually both ANA and anti-MPO antibodies. Further studies are needed to assess shared pathogenesis and therapeutic options. PMID:27258503

  16. Plasma exchange and glucocorticoid dosing in the treatment of anti-neutrophil cytoplasm antibody associated vasculitis (PEXIVAS): protocol for a randomized controlled trial

    OpenAIRE

    Walsh, Michael; Merkel, Peter A.; Peh, Chen Au; Szpirt, Wladimir; Guillevin, Loïc; Charles D. Pusey; deZoysa, Janak; Ives, Natalie; Clark, William F.; Quillen, Karen; Winters, Jeffrey L.; Wheatley, Keith; Jayne, David

    2013-01-01

    Background Granulomatosis with polyangiitis (GPA, Wegener’s) and microscopic polyangiitis (MPA) are small vessel vasculitides collectively referred to as anti-neutrophil cytoplasm antibody-associated vasculitis (AAV). AAV is associated with high rates of morbidity and mortality due to uncontrolled disease and treatment toxicity. Small randomized trials suggest adjunctive plasma exchange may improve disease control, while observational evidence suggests that current oral glucocorticoid doses a...

  17. Anti-neutrophil cytoplasmic antibody-enriched IgG induces adhesion of human T lymphocytes to extracellular matrix proteins.

    Science.gov (United States)

    Tomer, Y; Lider, O; Gilburd, B; Hershkoviz, R; Meroni, P L; Wiik, A; Shoenfeld, Y

    1997-06-01

    Recent studies have shown that anti-neutrophil cytoplasmic antibodies (ANCA) can activate neutrophils to adhere to endothelium, degranulate, and cause endothelial cell injury. These data have lead to the hypothesis that the T cell inflammatory response causing the vasculitis in Wegener's granulomatosis (WG) is secondary to stimulation of neutrophils by ANCA. So far there is no evidence for a direct effect of ANCA on lymphocytes. The present study was designed to examine whether lymphocytes can be directly stimulated by ANCA to adhere to endothelial extracellular matrix (ECM) proteins. Human and mouse ANCA-enriched IgG were tested for their ability to increase adhesion of human T lymphocytes to fibronectin, laminin, and intact ECM. Incubation of human T lymphocytes with human ANCA-enriched IgG increased adhesion of the lymphocytes in a dose-dependent manner to fibronectin, laminin, and intact ECM (the percentage adhesion to intact ECM was 55.7 +/- 3.1 and 45.0 +/- 1.0% for lymphocytes incubated with human IgG containing ANCA or control human IgG, respectively; P = 0.0045). The same induction of adhesion to fibronectin, laminin, and intact ECM was observed when the cells were incubated with the F(ab)2 fragment of ANCA-enriched IgG. Similarly, ANCA-enriched IgG produced in mice increased the adhesion of lymphocytes to fibronectin (the percentage adhesion to fibronectin was 29.7 +/- 4.3 and 16.6 +/- 1.9% for lymphocytes incubated with mouse IgG-ANCA or control mouse IgG, respectively; P = 0.0008). These results may suggest that ANCA can directly stimulate lymphocytes to adhere to endothelial ECM and to induce the vasculitic lesions of WG. It remains to be shown by which mechanisms ANCA stimulate lymphocytes to adhere to ECM. PMID:9175913

  18. Antineutrophil Cytoplasmic Antibodies Testing in a Large Cohort of Unselected Greek Patients

    Directory of Open Access Journals (Sweden)

    Konstantinos Tsiveriotis

    2011-01-01

    Full Text Available Objective. To retrospectively evaluate ANCA testing in a cohort of unselected Greek in- and outpatients. Methods. In 10803 consecutive serum samples, ANCA were tested by indirect immunofluorescence (IIF and ELISA. ELISA in inpatients was performed only on IIF positive sera. Results. Low prevalence (6.0% of IIF positive samples was observed. Among these samples, 63.5% presented perinuclear (p-ANCA, 9.3% cytoplasmic (c-ANCA and 27.2% atypical (x-ANCA pattern. 16.1% of p-ANCA were antimyeloperoxidase (anti-MPO positive, whereas 68.3% of c-ANCA were antiproteinase-3 (anti-PR3 positive. Only 17 IIF negative outpatients' samples were ELISA positive. ANCA-associated vasculitides (AAV, connective tissue disorders and gastrointestinal disorders represented 20.5%, 23.9%, and 21.2% of positive results, respectively. AAV patients exhibited higher rates of MPO/PR3 specificity compared to non-AAV (93.8% versus 8%. Conclusions. This first paper on Greek patients supports that screening for ANCA by IIF and confirming positive results by ELISA minimize laboratory charges without sacrificing diagnostic accuracy.

  19. Atualização do tratamento das vasculites associadas a anticorpo anticitoplasma de neutrófilos Treatment of antineutrophil cytoplasmic antibody-associated vasculitis: update

    OpenAIRE

    Alfredo Nicodemos Cruz Santana; Viktoria Woronik; Ari Stiel Radu Halpern; Barbas, Carmen S. V.

    2011-01-01

    As vasculites antineutrophil cytoplasmic antibody (ANCA, anticorpo anticitoplasma de neutrófilos) associadas (VAAs) são caracterizadas por uma inflamação sistêmica das artérias de pequeno e médio calibre (especialmente no trato respiratório superior e inferior, e nos rins). As VAAs compreendem a granulomatose de Wegener (agora chamada de granulomatose com poliangeíte), poliangeíte microscópica, VAA limitada ao rim e a síndrome de Churg-Strauss. Neste artigo, discutiremos as fases de tratament...

  20. Atualização do tratamento das vasculites associadas a anticorpo anticitoplasma de neutrófilos Treatment of antineutrophil cytoplasmic antibody-associated vasculitis: update

    Directory of Open Access Journals (Sweden)

    Alfredo Nicodemos Cruz Santana

    2011-12-01

    Full Text Available As vasculites antineutrophil cytoplasmic antibody (ANCA, anticorpo anticitoplasma de neutrófilos associadas (VAAs são caracterizadas por uma inflamação sistêmica das artérias de pequeno e médio calibre (especialmente no trato respiratório superior e inferior, e nos rins. As VAAs compreendem a granulomatose de Wegener (agora chamada de granulomatose com poliangeíte, poliangeíte microscópica, VAA limitada ao rim e a síndrome de Churg-Strauss. Neste artigo, discutiremos as fases de tratamento dessas vasculites, como fase de indução (com ciclofosfamida ou rituximab e fase de manutenção (com azatioprina, metotrexato ou rituximab. Além disso, discutiremos como manusear os casos refratários à ciclofosfamida.In its various forms, antineutrophil cytoplasmic antibody (ANCA-associated vasculitis (AAV is characterized by a systemic inflammation of the small and medium-sized arteries (especially in the upper and lower respiratory tracts, as well as in the kidneys. The forms of AAV comprise Wegener's granulomatosis (now called granulomatosis with polyangiitis, microscopic polyangiitis, renal AAV, and Churg-Strauss syndrome. In this paper, we discuss the phases of AAV treatment, including the induction phase (with cyclophosphamide or rituximab and the maintenance phase (with azathioprine, methotrexate, or rituximab. We also discuss how to handle patients who are refractory to cyclophosphamide.

  1. The antineutrophil antibody in uveitis.

    OpenAIRE

    Young, D W

    1991-01-01

    Ninety eight patients with uveitis of various types were tested for the presence of the antineutrophil antibody or ANCA by an indirect immunofluorescence method. This antibody is found in patients with diseases associated with small vessel vasculitis, including Wegener's granulomatosis and microscopic polyarteritis. Eleven true positive cases were found. A positive test was not associated with the anatomical site of the uveitis but was related to the time course of the disease. In particular ...

  2. False-positive myeloperoxidase binding activity due to DNA/anti-DNA antibody complexes: a source for analytical error in serologic evaluation of anti-neutrophil cytoplasmic autoantibodies.

    Science.gov (United States)

    Jethwa, H S; Nachman, P H; Falk, R J; Jennette, J C

    2000-09-01

    Anti-myeloperoxidase antibodies (anti-MPO) are a major type of anti-neutrophil cytoplasmic antibody (ANCA). While evaluating anti-MPO monoclonal antibodies from SCG/Kj mice, we observed several hybridomas that appeared to react with both MPO and DNA. Sera from some patients with systemic lupus erythematosus (SLE) also react with MPO and DNA. We hypothesized that the MPO binding activity is a false-positive result due to the binding of DNA, contained within the antigen binding site of anti-DNA antibodies, to the cationic MPO. Antibodies from tissue culture supernatants from 'dual reactive' hybridomas were purified under high-salt conditions (3 M NaCl) to remove any antigen bound to antibody. The MPO and DNA binding activity were measured by ELISA. The MPO binding activity was completely abrogated while the DNA binding activity remained. The MPO binding activity was restored, in a dose-dependent manner, by the addition of increasing amount of calf-thymus DNA (CT-DNA) to the purified antibody. Sera from six patients with SLE that reacted with both MPO and DNA were treated with DNase and showed a decrease in MPO binding activity compared with untreated samples. MPO binding activity was observed when CT-DNA was added to sera from SLE patients that initially reacted with DNA but not with MPO. These results suggest that the DNA contained within the antigen binding site of anti-DNA antibodies could bind to the highly cationic MPO used as substrate antigen in immunoassays, resulting in a false-positive test.

  3. Antineutrophil cytoplasmic autoantibodies in ulcerative colitis

    Directory of Open Access Journals (Sweden)

    Vodjgani M

    2000-10-01

    Full Text Available Antineutrophil cytoplasmic autoanibodies (ANCA were detecte in patients with certain autoimmune vascular disease such as Wegner’s granulomatosis, polyarthrits nodosa and systemic luuc erythematous. Indirect immunofluorescence (IIF technique was employed to detec these autoantibodies.ANCA have been recently detected in some forms of inflammatory bowel disease (IBD, ulcerative colitis (U.C. Crohn’s disease (C.D and primary sclerosing cholangitis (PSC. By IIF method, two general patterns of ANCA were seen: a cytoplasmic (C-ANCA and perinuclear form (P-ANCA. In this study we evaluated the presece of ANCA in 52 U.C. patients and 69 matched normal control group by IIF technique, and it’s relationship with disease activity. Site of colon involvement and, lesion extent. The results showed that all control group were ANCA negative, but 58% of patients had ANCA, and most cases (70% had C- ANCA. The obtained results also revealed that there was no relationship between ANCA and disease activity

  4. Anti-neutrophil cytoplasm autoantibodies (ANCA). The need for specific and sensitive assays

    DEFF Research Database (Denmark)

    Baslund, B; Petersen, J

    1998-01-01

    Anti-neutrophil cytoplasm antibodies (ANCA) are a group of autoantibodies primarily associated with systemic vasculitis. Hitherto, the method of choice for ANCA detection has been indirect immunofluorescence (IIF). By this method two major patterns can be seen: a cytoplasmic pattern (cANCA) or a...

  5. A case of myeloperoxidase anti-neutrophil cytoplasmic antibody (MPO-ANCA)-associated hypertrophic pachymeningitis presenting with multiple cranial nerve palsies and diabetes insipidus.

    Science.gov (United States)

    Yasuda, Ken; Sainouchi, Makoto; Goto, Masahiro; Murase, Nagako; Ohtani, Ryo; Nakamura, Michikazu

    2016-05-31

    A 61-year-old woman developed hearing difficulties and became thirsty after experiencing cold symptoms. A neurological examination revealed a loss of odor sensation, facial palsy, dysphasia, and dysarthria. Vocal cord palsy was observed during pharyngoscopy. Brain magnetic resonance imaging (MRI) showed a thickened pituitary stalk and swelling of the pituitary gland, but no high signal intensity regions were seen in the posterior portion of the pituitary gland. Gadolinium-enhanced MRI demonstrated a thickened dura mater over the anterior cranial fossa. A biopsy specimen of the thickened dura mater showed fibrosis, granulomatous inflammation, and necrotic foci. Blood tests detected myeloperoxidase anti-neutrophil cytoplasmic antibody (MPO-ANCA). The patient's urine osmolarity was low even though she exhibited hypernatremia. We diagnosed her with hypertrophic pachymeningitis associated with MPO-ANCA and diabetes insipidus. The patient received two courses of 5-day high-dose intravenous methylprednisolone (1.0 g/day), and was subsequently administered oral prednisolone, which gradually relieved her symptoms. However, the patient's symptoms recurred despite the high-dose prednisolone treatment. It was difficult to control the patient's symptoms in this case with oral prednisolone monotherapy, but combined treatment with cyclosporine resulted in sustained remission. It is considered that patients with MPO-ANCA-positive hypertrophic pachymeningitis require combination therapy with prednisolone and immunosuppressive agents at an early stage. PMID:27098904

  6. Experimental Autoimmune Vasculitis : An Animal Model of Anti-neutrophil Cytoplasmic Autoantibody-Associated Systemic Vasculitis

    OpenAIRE

    Little, Mark A.; Smyth, Lucy; Salama, Alan D; Mukherjee, Sriparna; Smith, Jennifer; Haskard, Dorian; Nourshargh, Sussan; Cook, H. Terence; Charles D. Pusey

    2009-01-01

    The morbidity burden associated with anti-neutrophil cytoplasmic autoantibody-associated vasculitis is increasing, and many novel biological therapies are now entering the drug development pipeline. There is thus an urgent need to develop a representative animal model to facilitate testing of these agents. We previously examined the effect of antineutrophil cytoplasmic autoantibody on leukocyte-endothelial interactions in WKY rats via immunization with human myeloperoxidase. We now seek to ex...

  7. Urinary matrix metalloproteinases reflect renal damage in anti-neutrophil cytoplasm autoantibody-associated vasculitis

    NARCIS (Netherlands)

    Sanders, J.S.F.; Huitema, M.G.; Hanemaaijer, R.; Goor, H. van; Kallenberg, C.G.M.; Stegeman, C.A.

    2007-01-01

    Renal expression of MMP-2, -9, and tissue inhibitor of MMP-1 (TIMP-1) correlates with histological disease activity in anti-neutrophil cytoplasm autoantibody (ANCA)-associated vasculitis (AAV). We studied whether urinary and plasma levels of MMP-2, -9, and TIMP-1 reflect renal expression of these pr

  8. Anti-neutrophil cytoplasmic autoantibodies and leukocyte-endothelial interactions : a sticky connection?

    NARCIS (Netherlands)

    Heeringa, P; Huugen, D; Tervaert, JWC

    2005-01-01

    Anti-neutrophil cytoplasmic autoantibodies (ANCA) with specificity for myeloperoxidase (MPO) or proteinase 3 (Pr3) are associated with systemic small-vessel vasculitides (SVV). Detection of ANCA is an established clinical tool in disease diagnosis and monitoring. Based on clinical and in vitro exper

  9. Current and novel biomarkers in anti-neutrophil cytoplasm-associated vasculitis.

    Science.gov (United States)

    Draibe, Juliana Bordignon; Fulladosa, Xavier; Cruzado, Josep Maria; Torras, Joan; Salama, Alan David

    2016-08-01

    Anti-neutrophil cytoplasm antibody (ANCA)-associated vasculitis (AAV) is characterized by a variable disease course, with up to 50% of patients having one relapse within 5 years and many progressing to end-stage organ damage despite modern treatment strategies. Moreover, complications arising from treatment dominate the causes of mortality and morbidity both early and late during disease, especially in the elderly and those with severe renal involvement, and there is additional uncertainty as to how long treatment should be continued. There is, therefore, an urgent clinical need to identify robust biomarkers to better predict treatment responses, risk of disease relapse and eventual complete clinical and immunological quiescence. To date, no such biomarkers exist, but better understanding of disease pathogenesis and the underlying immune dysfunction has provided some potential candidates linked to the discovery of new antibodies, different leukocyte activation states, the role of the alternative complement pathway and markers of vascular activation. With all promising new biomarkers, there is the need to rapidly replicate and validate early findings using large biobanks of samples that could be brought together by leaders in the field.

  10. Complement Activation Is Involved in Renal Damage in Human Antineutrophil Cytoplasmic Autoantibody Associated Pauci-Immune Vasculitis

    NARCIS (Netherlands)

    Xing, Guang-qun; Chen, Min; Liu, Gang; Heeringa, Peter; Zhang, Jun-jun; Zheng, Xin; Jie, E.; Kallenberg, Cees G. M.; Zhao, Ming-hui

    2009-01-01

    This study was to investigate the evidence for complement activation in renal biopsy specimens of patients with myeloperoxidase (MPO)-antineutrophil cytoplasmic autoantibody (ANCA)-associated pauci-immune vasculitis. Renal biopsy specimens from seven patients with MPO-ANCA positive pauci-immune necr

  11. Inflammatory Bowel Disease in Indo-Canadians with and without Antineutrophil Cytoplasmic Autoantibodies

    Directory of Open Access Journals (Sweden)

    Hugh James Freeman

    2000-01-01

    Full Text Available A sequentially evaluated cohort of Indo-Canadians with either ulcerative colitis or Crohn’s disease were prospectively examined for antineutrophil cytoplasmic autoantibodies (ANCA. Of 84 patients, 62 had ulcerative colitis and 22 had Crohn’s disease. About one-third were born in Canada, and two-thirds were migrants from India or other countries, particularly East African nations. There was a disease-based and geographically based male predominance. The mean age of Canadian-born patients was significantly less than that of those born in other countries. Moreover, for migrants, the mean duration of residence in Canada before developing disease was 8.9 years for Crohn’s disease patients and 13.5 years for ulcerative colitis patients. Moderate to severe disease was present; virtually all those with Crohn’s disease had colonic involvement, and most of those with ulcerative colitis had extensive colonic disease. Overall, 40 of 84 (48% were seropositive for ANCA, including a majority of those with ulcerative colitis but not Crohn’s disease. In addition, eight had cytoplasmic ANCA, a reported seromarker for extensive colitis. Seropositive and seronegative patients were similar in age, sex, birth or duration of residence in Canada, site and severity of disease, familial history and complications, including pouchitis. This study supports the view that these diseases arise in individuals with a genetic predisposition following exposure to some, as yet unknown, environmental factor.

  12. Urinary levels of high mobility group box-1 are associated with disease activity in antineutrophil cytoplasmic autoantibody-associated vasculitis.

    Directory of Open Access Journals (Sweden)

    Tian-Tian Ma

    Full Text Available High mobility group box-1 (HMGB1, a kind of pro-inflammatory mediator, is associated with inflammatory conditions and tissue damage. Our previous study demonstrated that the circulating levels of HMGB1 correlated with disease activity of antineutrophil cytoplasmic antibody (ANCA-associated vasculitis (AAV. In the current study, we aimed to measure urinary levels of HMGB1 in AAV patients, correlated them to clinical activity index and analysed the immunohistochemical HMGB1 staining in kidney specimens.50 patients with AAV in active stage and 56 patients with AAV in remission were recruited. The urinary levels of HMGB1 were determined by enzyme-linked immunosorbent assay. Moreover, renal biopsy specimens from 27 patients with active AAV were randomly collected to evaluate the deposition of HMGB1.Urinary HMGB1 levels in AAV patients in active stage were significantly higher than those in AAV patients in remission and healthy controls (1.46 [0.56-3.43] versus 0.38 [0.10-1.35] mg/μmolCr, P=0.001; 1.46 [0.56-3.43] versus 0.48 [0.40-0.60] mg/μmolCr, P=0.000, respectively. Further analysis found that urinary levels of HMGB1 correlated with erythrocyte sedimentation rate (r=0.354, p=0.012, C-reactive protein (r=0.289, p=0.042, and Birmingham Vasculitis Activity Score (r=0.350, p=0.013. Renal tissue of active AAV patients showed HMGB1 was mainly expressed in the cytoplasm and the extracellular space. The percentage of HMGB1-negative nuclei in renal tissue of patients with active AAV was significantly higher than that in normal controls (60.6±20.2 % versus 2.7±0.6 %, p<0.01.Urinary levels of HMGB1 may be associated with the disease activity in AAV patients.

  13. ANCA在系统性红斑狼疮肺栓塞中的诊断价值%Diagnostic value of antineutrophil cytoplasmic antibodies in patients with systemic lupus erythematosus merge pulmonary embolism

    Institute of Scientific and Technical Information of China (English)

    杨栋; 宫怡; 杨吉娟

    2009-01-01

    目的 观察抗中性粒细胞胞浆抗体(Anti-neutrophil cytopasmic antibodies,ANCA)在系统性红斑狼疮(Systematic lupus erythematosus,SLE)合并肺栓塞(Pulmonary Embolism,PE)诊断中的价值.方法 比较36例系统性红斑狼疮肺栓塞患者与50例无肺栓塞的系统性红斑狼疮患者的临床表现及ANCA阳性率.结果 系统性红斑狼疮肺栓塞患者组ANCA阳性率明显高于无肺栓塞的系统性红斑狼疮患者组.结论 血清ANCA抗体有助于系统性红斑狼疮肺栓塞的诊断.

  14. Exploration, Development, and Validation of Patient-reported Outcomes in Antineutrophil Cytoplasmic Antibody–associated Vasculitis Using the OMERACT Process

    Science.gov (United States)

    Robson, Joanna C.; Milman, Nataliya; Tomasson, Gunnar; Dawson, Jill; Cronholm, Peter F.; Kellom, Katherine; Shea, Judy; Ashdown, Susan; Boers, Maarten; Boonen, Annelies; Casey, George C.; Farrar, John T.; Gebhart, Don; Krischer, Jeffrey; Lanier, Georgia; McAlear, Carol A.; Peck, Jacqueline; Sreih, Antoine G.; Tugwell, Peter; Luqmani, Raashid A.; Merkel, Peter A.

    2016-01-01

    Objective Antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) is a group of linked multisystem life- and organ-threatening diseases. The Outcome Measures in Rheumatology (OMERACT) vasculitis working group has been at the forefront of outcome development in the field and has achieved OMERACT endorsement of a core set of outcomes for AAV. Patients with AAV report as important some manifestations of disease not routinely collected through physician-completed outcome tools; and they rate common manifestations differently from investigators. The core set includes the domain of patient-reported outcomes (PRO). However, PRO currently used in clinical trials of AAV do not fully characterize patients’ perspectives on their burden of disease. The OMERACT vasculitis working group is addressing the unmet needs for PRO in AAV. Methods Current activities of the working group include (1) evaluating the feasibility and construct validity of instruments within the PROMIS (Patient-Reported Outcome Measurement Information System) to record components of the disease experience among patients with AAV; (2) creating a disease-specific PRO measure for AAV; and (3) applying The International Classification of Functioning, Disability and Health to examine the scope of outcome measures used in AAV. Results The working group has developed a comprehensive research strategy, organized an investigative team, included patient research partners, obtained peer-reviewed funding, and is using a considerable research infrastructure to complete these interrelated projects to develop evidence-based validated outcome instruments that meet the OMERACT filter of truth, discrimination, and feasibility. Conclusion The OMERACT vasculitis working group is on schedule to achieve its goals of developing validated PRO for use in clinical trials of AAV. (First Release September 1 2015; J Rheumatol 2015;42:2204–9; doi:10.3899/jrheum.141143) PMID:26329344

  15. Serum interleukin-1 8 level in patients with antineutrophil cytoplasmic antibodies associated vasculitis and implication%抗中性粒细胞胞浆抗体相关性血管炎患者白细胞介素18的水平及意义

    Institute of Scientific and Technical Information of China (English)

    蔡娟; 王卫黎; 黄鑫; 王渊; 唐莎; 张静波

    2015-01-01

    目的:检测抗中性粒细胞胞浆抗体相关性血管炎(antineutrophil cytoplasmic antibodies associated vasculitis,AAV)患者白细胞介素18(interleukin-18,IL-18)的水平,探讨 IL-18在 AAV 发病中的意义。方法选择2013年4月至2014年7月在第三军医大学附属新桥医院肾内科住院诊治的 AAV 患者27例设为 AAV 组,选择17例有蛋白尿、镜下血尿且肾活检明确诊断为轻度系膜增生性肾小球肾炎(mesangial proliferative glomerulonephritis,MsPGN)患者设为 MsPGN 组,另外选择17名健康者设为健康对照组,收集3组血清及其临床资料,采用酶联免疫吸附试验(enzymelinked im-munosorbent assay,ELISA)方法检测3组患者血清 IL-18浓度;Pearson 积矩相关分析 AAV 患者血清 IL-18水平与 AAV 活动性指标伯明翰血管炎活动性评分(birmingham vasculitis active scoring sys-tem,BVAS)、C 反应蛋白(C-reaction protein,CRP)水平、红细胞沉降率(erythrocyte sedimentation rate,ESR)、估算肾小球滤过率(estimated glomerular filtration rate,eGFR)、血肌酐(SCr)水平及24 h尿蛋白定量的相关性;并对活动期和缓解期 AAV 患者的血清 IL-18、CRP 和 ESR 水平行受试者工作特征(receiver operating characteristic,ROC)曲线的曲线下面积(area under the curve,AUC)评价 IL-18作为评估疾病活动性指标的准确性。结果①活动期 AAV 患者 IL-18浓度[(183.85±57.0)ng/L]高于缓解期 AAV 患者 IL-18水平[(92.0±34.29)ng/L](P 0.05)无明显相关;③ROC 曲线分析显示,相比血清 CRP 和 ESR 水平,IL-18水平能更好地区分 AAV 患者的活动性。结论AAV 患者活动期 IL-18水平明显升高,且与疾病的活动性密切相关,提示 IL-18可能参与 AAV 的病理损伤过程。%Objective To investigate serum interleukin-1 8(IL-1 8)level in patients with antineu-trophil cytoplasmic antibodies(ANCA)associated vasculitis(AAV)and the implication in AAV.Meth-ods Sera and clinical data were obtained from

  16. Anticorpo anticitoplasma de neutrófilos (ANCA em pioderma gangrenoso, um marcador sorológico para associação com doenças sistêmicas: estudo de oito casos Antineutrophil cytoplasmic antibody (ANCA in pyoderma gangrenosum, a serologic marker for associated systemic diseases: a study of eight cases

    Directory of Open Access Journals (Sweden)

    Virgínia Lúcia Ribeiro Cabral

    2004-02-01

    Full Text Available FUNDAMENTOS: A etiopatogenia da retocolite ulcerativa inespecífica (RCUI e de suas manifestações extra-intestinais permanece em discussão, embora o envolvimento do sistema imune seja enfatizado, e uma possível participação dos neutrófilos é demonstrada pela detecção do anticorpo anticitoplasma de neutrófilo (ANCA nessa doença inflamatória intestinal. O pioderma gangrenoso (PG é considerado manifestação cutânea rara da retocolite ulcerativa, e o Anca também tem sido detectado nessa dermatose. OBJETIVOS: Investigar a relação entre o comportamento clínico da RCUI e o aparecimento do PG e sua associação com ANCA. CASUÍSTICA E MÉTODOS: Anca foi pesquisado nos soros de oito pacientes com PG, quatro apresentando RCUI, e os outros, PG não associado a doenças sistêmicas. RESULTADOS: Não se detectou o Anca nos soros dos portadores exclusivamente de pioderma gangrenoso. Dois casos de pancolite em atividade inflamatória acompanhada de pioderma e colangite esclerosante primária (CEP apresentaram positividade para ANCA, enquanto os soros de dois outros pacientes com RCUI e PG tiveram resultados negativos. CONCLUSÕES: A Presença de ANCA nos soros de pacientes com PG associado a RCUI e CEP sugere que a associação com CEP seja responsável pela positividade do ANCA na presente amostra.BACKGROUND: The pathogenesis of Ulcerative Colitis (UC and its extraintestinal manifestations remain uncertain, although involvement of the immune system is emphasized. The likely importance of neutrophils is demonstrated by detection of the antineutrophil cytoplasmic antibody (ANCA in this inflammatory bowel disease. Pyoderma Gangrenosum (PG is an idiopathic skin condition and a rare cutaneous manifestation of UC. ANCA has also been reported in the latter dermatosis. OBJECTIVES: To invetigate the relationship between clinical features of UC and the appearance of PG and its association with ANCA. PATIENTS AND METHODS: ANCA was determined in sera

  17. Clinical Characteristis of Primary Antineutrophil Cytoplasmic Antibody-associated Vasculitis with Diffuse Interstitial Lung Diseases%原发性抗中性粒细胞胞浆抗体相关性血管炎伴弥漫性间质性肺疾病临床特点分析

    Institute of Scientific and Technical Information of China (English)

    孙越; 方秋红; 马迎民; 原庆; 黄爱本

    2013-01-01

    目的 探讨原发性抗中性粒细胞胞浆抗体(ANCA)相关性血管炎(AASV)伴弥漫性间质性肺疾病(DILD)的临床特点.方法 回顾性分析42例AASV-DILD患者的临床资料,将上述患者分为以呼吸道症状及非呼吸道症状为主要临床表现的A组和B组,对两组资料进行比较.结果 (1)A组24例,B组18例,临床症状两组相比咳嗽(17/24 比 1/18)、气短(12/24 比 0)、蛋白尿血尿并水肿(3/24 比 14/18)、关节肌肉疼痛(1/24 比 6/18)发生率差异均有统计学意义(P<0.05).(2)A、B两组年龄、病程、血气分析、肺功能检查、支气管镜检查、抗核抗体滴度、ANCA阳性率及类别差异均无统计学意义(P>0.05).(3)30例患者发生肾功能衰竭(RF),急性肾功能衰竭(ARF)发生率为14/42;慢性肾功能衰竭(CRF)发生率B组为11/18,高于A组发生率5/24,差异有统计学意义(P<0.05).(4)8例患者超声心动图提示肺心病(PHD)表现,A组发生率(7/13)高于B组(1/11),差异有统计学意义(P<0.05).(5)8例患者行经支气管镜肺活检(TBLB),2例提示小血管炎病理改变;11例支气管肺泡灌洗液(BALF)提示中性粒细胞百分比增高.(6)33例行高分辨CT(HRCT)检查,主要影像学表现为网格影、蜂窝肺、磨玻璃影、小叶间隔增厚,磨玻璃影两组发生率(5/21 比 9/12)差异有统计学意义(P<0.05).结论 (1)AASV-DILD患者表现为多脏器受累,当以非呼吸系统症状为主要表现时,呼吸系统病变易漏诊.(2)AASV-DILD除肺脏外,肾脏是最易受累的脏器,有较高的ARF发生率,CRF多见于以肾脏受累为主要临床症状患者.(3)PHD多发生于以呼吸系统症状为主的患者.(4)AASV-DILD患者BALF 以中性粒细胞增高为主,TBLB在诊断AASV中有一定价值.(5)AASV-DILD影像学无特征性,应常规进行ANCA检测,避免漏诊.%Objective To explore the features of primary antineutrophil cytoplasmic antibody - associated vasculitis with diffuse interstitial lung diseases

  18. 抗中性粒细胞胞质抗体检测在风湿病诊断中的临床价值探讨%A study on clinical values of antineutrophil cytoplasmic antibody detection in diagnosis of rheumatic diseases

    Institute of Scientific and Technical Information of China (English)

    石莉萍; 张利方; 吴颖涛; 秦滢

    2014-01-01

    目的:探讨抗中性粒细胞胞质抗体(ANCA)检测对风湿病患者诊断的临床意义。方法将965例风湿病患者作为风湿病组,其中,SLE 256例,MCTD 124例,RA 336例,干燥综合征(SS)166例,系统性硬化症(SSc)45例,皮肌炎38例。将400例健康体检者作为对照组。采用间接免疫荧光法(IIF)检测胞质型ANCA(c-ANCA)、核周型ANCA (p-ANCA)。ANCA阳性者采用酶联免疫吸附测定(ELISA)检测其抗髓过氧化物酶(MPO)抗体。结果风湿病组、对照组受检者血清ANCA的阳性率的差异有统计学意义(P<0.01),两组受检者ANCA阳性的血清中,p-ANCA与c-ANCA的阳性率的差异有统计学意义(P<0.01)。伴SLE的患者血清p-ANCA的阳性率显著高于伴MCTD、RA、SS、SSc、皮肌炎患者及对照组受检者(P<0.05);伴MCTD的患者血清p-ANCA的阳性率显著高于伴RA、SS、SSc、皮肌炎者及对照组受检者(P<0.05)。165例p-ANCA阳性的风湿病组患者抗MPO抗体的阳性率为58.4%,显著高于对照组受检者(3%)(P<0.01);ANCA阳性的风湿病组患者伴肾功能损害的阳性率为55.4%,显著高于ANCA阴性的风湿病组患者(21.8%)及对照组受检者(6.25%)(P<0.01)。结论血清ANCA检测对预防风湿病患者肾毒性损害有重要意义。%Objective To investigate the clinical values of antineutrophil cytoplasmic antibody (ANCA) detection in the diagnosis of rheumatic diseases .Methods 965 patients with rheumatic diseases were taken as rheumatic group ,including 256 cases of SLE , 124 of mixed connective tissue disease(MCTD) ,336 of rheumatoid arthritis(RA) ,166 of Sjogren′s Syndrome(SS) ,45 of systemic sclerosis(SSc) and 38 of dermatomyositis .400 healthy people were served as the control group .Indirect immunofluorescence (IIF) was employed to detect the cytoplasmic ANCA (c-ANCA ) ,perinuclear ANCA (p

  19. Serum soluble interleukin 2 receptor levels in anti-neutrophil cytoplasmic autoantibodies--positive systemic vasculitis.

    OpenAIRE

    Lai, K N; Lockwood, C. M.

    1993-01-01

    Systemic vasculitis is characterized by the presence of autoantibodies to neutrophil cytoplasmic antigens (ANCA). The role of T-lymphocytes in systemic vasculitis remains uncertain. In the present study, we attempted to explore the role of T-lymphocytes in systemic vasculitis by measuring the serum soluble interleukin 2 receptor (sIL2R) levels in seven vasculitic patients and comparing the sequential measurements with the titres of ANCA which satisfactorily reflect the disease activity. The s...

  20. 钙卫蛋白及抗中性粒细胞胞浆抗体联合检测对炎症性肠病的诊断价值研究%Diagnostic value of combined detection of calprotectin and antineutrophil cytoplasmic antibodies in inflammatory bowel disease

    Institute of Scientific and Technical Information of China (English)

    韩晓芳; 刘洋; 谭艳; 杨国香; 贾海琴

    2012-01-01

    目的 探讨钙卫蛋白与抗中性粒细胞胞浆抗体(ANCA)联合检测对炎症性肠病(IBD)的诊断价值.方法 收集确诊为IBD的患者79例作为IBD组,腹痛、腹泻等排除IBD的患者42例作为疾病对照组,健康体检者34例作为健康对照组.分别检测血液样本中ANCA和粪便样本中钙卫蛋白的水平.结果 79例IBD患者粪便钙卫蛋白浓度为(493.86±204.18)μg/g高于疾病对照组[(71.46±60.51)μg/g]和健康对照组[(36.19±13.46)μg/g].钙卫蛋白在IBD组、疾病对照组和健康对照组中的阳性率分别为57.0%、19.0%、0.0%;ANCA在三组中的阳性率分别为63.3%、4.8%、0.0%.IBD组的钙卫蛋白、ANCA阳性率显著高于其他两组(P<0.05).钙卫蛋白与ANCA联合检测在IBD组、疾病对照组和健康对照组中的阳性率分别为78.5%、23.8%和0.0%.结论 钙卫蛋白和ANCA联合检测可显著提高IBD的诊断率,为临床IBD的早期诊断和治疗提供可靠依据.%Objective To explore the diagnostic value of combined detection of calprotectin and antineutrophil cytoplasniic anti bodies(ANCA) in inflammatory bowel disease(IBD). Methods 79 patients with definite diagnosis of IBD(IBD group) ,42 patients with abdominal pain and diarrhea,but without IBD(disease control group) and 34 cases of healthy subjects(healthy control group) were enrolled,and detected for ANCA level in blood sample and calprotectin level in and fecal sample. Results The fecal calprotec tin concentration was (493. 86 + 204. 18) μg/g in IBD group, higher than (71. 46+60. 51) and (36. 19 + 13. 46) μg/g in disease control group and healthy control group, respectively. The positive rates of calprotectin in IBD group, disease control group and healthy control group were 57. 0% ,19. 0% and 0. 0%,of ANCA in the three groups were 63. 3%,4. 8% and 0. 0%. The positive rates of calprotectin and ANCA in IBD group were higher than those in the other two groups(P<0. 05). The positive rates of com bined detection of

  1. Anti-proteinase 3 anti-neutrophil cytoplasm autoantibodies recapitulate systemic vasculitis in mice with a humanized immune system.

    Directory of Open Access Journals (Sweden)

    Mark A Little

    Full Text Available Evidence is lacking for direct pathogenicity of human anti-proteinase-3 (PR3 antibodies in development of systemic vasculitis and granulomatosis with polyangiitis (GPA, Wegener's granulomatosis. Progress in study of these antibodies in rodents has been hampered by lack of PR3 expression on murine neutrophils, and by different Fc-receptor affinities for IgG across species. Therefore, we tested whether human anti-PR3 antibodies can induce acute vasculitis in mice with a human immune system. Chimeric mice were generated by injecting human haematopoietic stem cells into irradiated NOD-scid-IL2Rγ⁻/⁻ mice. Matched chimera mice were treated with human IgG from patients with: anti-PR3 positive renal and lung vasculitis; patients with non-vasculitic renal disease; or healthy controls. Six-days later, 39% of anti-PR3 treated mice had haematuria, compared with none of controls. There was punctate bleeding on the surface of lungs of anti-PR3 treated animals, with histological evidence of vasculitis and haemorrhage. Anti-PR3 treated mice had mild pauci-immune proliferative glomerulonephritis, with infiltration of human and mouse leukocytes. In 3 mice (17% more severe glomerular injury was present. There were no glomerular changes in controls. Human IgG from patients with anti-PR3 autoantibodies is therefore pathogenic. This model of anti-PR3 antibody-mediated vasculitis may be useful in dissecting mechanisms of microvascular injury.

  2. Anti-proteinase 3 anti-neutrophil cytoplasm autoantibodies recapitulate systemic vasculitis in mice with a humanized immune system.

    LENUS (Irish Health Repository)

    Little, Mark A

    2012-01-01

    Evidence is lacking for direct pathogenicity of human anti-proteinase-3 (PR3) antibodies in development of systemic vasculitis and granulomatosis with polyangiitis (GPA, Wegener\\'s granulomatosis). Progress in study of these antibodies in rodents has been hampered by lack of PR3 expression on murine neutrophils, and by different Fc-receptor affinities for IgG across species. Therefore, we tested whether human anti-PR3 antibodies can induce acute vasculitis in mice with a human immune system. Chimeric mice were generated by injecting human haematopoietic stem cells into irradiated NOD-scid-IL2Rγ⁻\\/⁻ mice. Matched chimera mice were treated with human IgG from patients with: anti-PR3 positive renal and lung vasculitis; patients with non-vasculitic renal disease; or healthy controls. Six-days later, 39% of anti-PR3 treated mice had haematuria, compared with none of controls. There was punctate bleeding on the surface of lungs of anti-PR3 treated animals, with histological evidence of vasculitis and haemorrhage. Anti-PR3 treated mice had mild pauci-immune proliferative glomerulonephritis, with infiltration of human and mouse leukocytes. In 3 mice (17%) more severe glomerular injury was present. There were no glomerular changes in controls. Human IgG from patients with anti-PR3 autoantibodies is therefore pathogenic. This model of anti-PR3 antibody-mediated vasculitis may be useful in dissecting mechanisms of microvascular injury.

  3. Update on Anti-Saccharomyces cerevisiae antibodies, anti-nuclear associated anti-neutrophil antibodies and antibodies to exocrine pancreas detected by indirect immunofluorescence as biomarkers in chronic inflammatory bowel diseases: Results of a multicenter study

    Institute of Scientific and Technical Information of China (English)

    S Desplat-Jégo; JC Grimaud; M Veyrac; P Chamouard; RL Humbel; C Johanet; A Escande; J Goetz; N Fabien; N Olsson; E Ballot; J Sarles; JJ Baudon

    2007-01-01

    AIM:Anti-Saccharomyces cerevisiae antibodies (ASCA), anti-nuclear associated anti-neutrophil antibodies (NANA) and antibodies to exocrine pancreas (PAB), are serological tools for discriminating Crohn's disease (CrD) and ulcerative colitis (DC). Like CrD, coeliac disease (CoD) is an inflammatory bowel disease (IBD) associated with (auto) antibodies. Performing a multicenter study we primarily aimed to determine the performance of ASCA, NANA and PAB tests for IBD diagnosis in children and adults, and secondarily to evaluate the prevalence of these markers in CoD.METHODS: Sera of 109 patients with CrD, 78 with UC, 45 with CoD and 50 healthy blood donors were retrospectively included. ASCA, NANA and PAB were detected by indirect immunofluorescence (IIF).RESULTS: ASCA+/NANA- profile displayed a positive predictive value of 94.2% for CrD. Detection of ASCA was correlated with a more severe clinical profile of CrD and treatment of the disease did not influence their serum levels. ASCA positivity was found in 37.9% of active CoD.PAB were found in 36.7% CrD and 13.3% CoD patients and were not correlated with clinical features of CrD, except with an early onset of the disease. Fifteen CrD patients were ASCA negative and PAB positive.CONCLUSION: ASCA and PAB detected by IIF are specific markers for CrD although their presence does not rule out a possible active CoD. The combination of ASCA, NANA and PAB tests improves the sensitivity of immunological markers for CrD. Repeating ASCA, NANA, and PAB testing during the course of CrD has no clinical value.

  4. Perinuclear anti-neutrophil cytoplasmic antibodies (p-anca) in chronic ulcerative colitis: Experience in a Mexican institution

    Institute of Scientific and Technical Information of China (English)

    Jesus K Yamamoto-Furusho; Takeshi Takahashi-Monroy; Omar Vergara-Fernandez; Edgardo Reyes; Luis Uscanga

    2006-01-01

    AIM: To assess the prevalence and clinical value of p-ANCA in a sample of Mexican ulcerative colitis (UC) patients.METHODS: In a prospective, IRB-approved protocol,p-ANCA was determined in 80 patients with UC (mean age, 32 ± 12.9 years). The severity and extension of disease were determined by clinical methods, searching a statistical association with p-ANCA status.RESULTS: p-ANCA were detected in 41 (51%) patients.Severity of disease was the only clinical variable statistically associated with their presence (P < 0.0001; OR = 9;CI 95% = 3.2-24.7).CONCLUSION: The prevalence of p-ANCA was similar to that reported in other countries. Their presence was associated to UC severity, but offered no more information than the obtained by clinical methods.

  5. Distinction between antinuclear antibody and P-ANCA.

    OpenAIRE

    Lee, S. S.; Lawton, J W; Chak, W

    1991-01-01

    To differentiate between perinuclear immunofluorescence staining of antinuclear antibody (ANA) and the perinuclear form of anti-neutrophil cytoplasmic antibody (P-ANCA), the pattern after formaldehyde vapour fixation of normal human neutrophils was compared with that of standard ethanol fixation. Fifteen out of 17 myeloperoxidase antibody positive sera showed cytoplasmic staining on formaldehyde vapour fixed cells; 30 of the 32 ANA positive samples became negative or gave weak nuclear stainin...

  6. Association of the HLA-DRB1*0701 allele with perinuclear anti-neutrophil cytoplasmatic antibodies in Mexican patients with severe ulcerative colitis

    Institute of Scientific and Technical Information of China (English)

    Jesus K Yamamoto-Furusho; Luis Uscanga-Domínguez; Alondra Lopez-Martinez; Julio Granados

    2006-01-01

    AIM: To determine the association between the HLA-DRB1 alleles and perinuclear anti-neutrophil cytoplasmatic antibodies (p-ANCA) positive in Mexican patients with ulcerative colitis (UC).METHODS: Ninety Mexican mestizo patients (45 females) with UC, confirmed by biopsy, were studied. High resolution HLA typing was performed by PCR-SSO reverse dot blot and PCR-SSP. Molecular typing techniques were applied to define HLA-DRB1 alleles. Enzyme-linked immunosorbent assay and immunofluorescence techniques were used to detect p-ANCA.RESULTS: Forty-eight (53%) UC patients were positive for p-ANCA by ELISA and IF. We found that p-ANCA-positive UC patients had a significantly increased frequency of HLA-DR7 compared with p-ANCA-negative controls (22% vs 5.1%; pC=0.02, OR=5.2, CI 95%:1.06-37.82). Disease activity was scored as severe in 20 patients, moderate in 8, mild in 14 and no activity in the remaining 38 patients according to the Truelove and Witts criteria. Subgroup analysis showed a significantly increased frequency of the HLA-DRB1*07 allele in 15 of 20 UC patients with severe activity of UC and p-ANCA positivity [100% vs 0%; pC=0.0000001; OR=35]. No significant differences were found between p-ANCA positive patients, HLA-DR alleles and other clinical features such as extraintestinal manifestations, proctocolectomy and extension.CONCLUSION: The HLA-DRB1*07 is associated with p-ANCA positive UC Mexican patients.

  7. Analysis of a solid-phase radioimmunoassay for antibodies to cytoplasmic antigen fractions of Candida albicans

    International Nuclear Information System (INIS)

    An indirect solid-phase radioimmunoassay (SPRIA) in individual polystyrene microtiter cups has been adapted for measurement of antibody to various cytoplasmic and carbohydrate antigen fractions of Candida albicans. The assay was optimized for sensitivity, precision and linearization of serum dilution curves. The optimized procedure allows computerized measurement of anti-Candida antibodies and can be used for measurement of antibody over a wide concentration range. The procedure obviates variation due to changes in day-to-day counts as a result of isotope decay and end-point antibody dilutions. The assay has been used to demonstrate a Poisson-like distribution of antibody levels in the sera of persons showing no symptoms of candidiasis. The minimum antibody level detectable by the assay is about two orders of magnitude lower than the lowest level found in human serum and 4 orders of magnitude lower than the most sensitive test used hitherto, the hemagglutination test. (Auth.)

  8. Antibody-induced down-regulation of a mutated insulin receptor lacking an intact cytoplasmic domain

    International Nuclear Information System (INIS)

    Insulin receptor down-regulation was studied in various Chinese hamster ovary (CHO) cell lines expressing transfected human insulin receptor cDNAs. In addition to a cell line expressing the normal receptor (CHO.T line), three lines expressing mutated receptors were studied: the CHO.T-t line, which expresses a receptor with a degraded cytoplasmic domain due to the removal of the C-terminal 112 amino acids, and the CHO.YF1 and CHO.YF3 lines, in which important autophosphorylation sites of the receptor kinase (tyrosines-1162 and -1163) have been replaced by phenylalanine. A monoclonal anti-receptor antibody, but not insulin itself, was found to down-regulate cell surface receptor levels in all four cell lines by 60-80% after 18-h treatment at 370C. Down-regulation of the CHO.T and CHO.T-t receptors occurred at similar antibody concentrations and with a similar time course, although the maximum level of CHO.T-t down-regulation (60%) was generally lower than the level of CHO.T down-regulation (80%). Pulse-chase labeling of these two cell types with [35S]methionine revealed that antibody treatment of both CHO.T and CHO.T-t cells resulted in a similar increase in the rate of degradation of mature receptor subunits. These results indicate that antibody-induced down-regulation of the insulin receptor in these cells can occur in the absence of various autophosphorylation sites of the receptor and that the mechanism of antibody-induced down-regulation is different from that for insulin

  9. Evaluation of capture ELISA for detection of antineutrophil cytoplasmic antibodies directed against proteinase 3 in Wegener's granulomatosis : first results from a multicentre study

    NARCIS (Netherlands)

    Csernok, E; Holle, J; Hellmich, B; Willem, J; Tervaert, C; Kallenberg, CGM; Limburg, PC; Niles, J; Pan, GL; Specks, U; Westman, K; Wieslander, J; Gross, WL

    2004-01-01

    Objective: To evaluate the performance characteristics of direct and capture ELISA for the detection of PR3-ANCA in Wegener's granulomatosis (WG) in international ANCA reference laboratories. Methods: Serum samples were derived from patients with histological and clinical diagnosis of WG (n = 60), r

  10. Detection of Antibodies to Brucella Cytoplasmic Proteins in the Cerebrospinal Fluid of Patients with Neurobrucellosis

    Science.gov (United States)

    Baldi, Pablo C.; Araj, George F.; Racaro, Graciela C.; Wallach, Jorge C.; Fossati, Carlos A.

    1999-01-01

    The diagnosis of human neurobrucellosis usually relies on the detection of antibodies to Brucella lipopolysaccharide (LPS) in cerebrospinal fluid (CSF) by agglutination tests or enzyme-linked immunosorbent assay (ELISA). Here we describe the detection of immunoglobulin G (IgG) to cytoplasmic proteins (CP) of Brucella spp. by ELISA and Western blotting in seven CSF samples from five patients with neurobrucellosis. While IgG to CP (titers of 200 to 12,800) and IgG to LPS (800 to 6,400) were found in the CSF of these patients, these antibodies were not detected in CSF samples from two patients who had systemic brucellosis without neurological involvement. The latter, however, had serum IgG and IgM to both LPS and CP. No reactivity to these antigens was found in CSF samples from 14 and 20 patients suffering from nonbrucellar meningitis and noninfectious diseases, respectively. These findings suggest that, in addition to its usefulness in the serological diagnosis of human systemic brucellosis, the ELISA with CP antigen can be used for the specific diagnosis of human neurobrucellosis. PMID:10473531

  11. Expression of recombinant multi-coloured fluorescent antibodies in gor -/trxB- E. coli cytoplasm

    Directory of Open Access Journals (Sweden)

    Markiv Anatoliy

    2011-11-01

    Full Text Available Abstract Background Antibody-fluorophore conjugates are invaluable reagents used in contemporary molecular cell biology for imaging, cell sorting and tracking intracellular events. However they suffer in some cases from batch to batch variation, partial loss of binding and susceptibility to photo-bleaching. In theory, these issues can all be addressed by using recombinant antibody fused directly to genetically encoded fluorescent reporters. However, single-chain fragment variable domains linked by long flexible linkers are themselves prone to disassociation and aggregation, and in some cases with isoelectric points incompatible with use in physiologically relevant milieu. Here we describe a general approach that permits fully functional intracellular production of a range of coloured fluorescent recombinant antibodies with optimally orientated VH/VL interfaces and isoelectric points compatible for use in physiological solutions at pH 7.4 with a binding site to fluorophore stoichiometry of 1:1. Results Here we report the design, assembly, intracellular bacterial production and purification of a panel of novel antibody fluorescent protein fusion constructs. The insertion of monomeric fluorescent protein derived from either Discosoma or Aequorea in-between the variable regions of anti-p185HER2-ECD antibody 4D5-8 resulted in optimal VH/VL interface interactions to create soluble coloured antibodies each with a single binding site, with isoelectric points of 6.5- 6. The fluorescent antibodies used in cell staining studies with SK-BR-3 cells retained the fluorophore properties and antibody specificity functions, whereas the conventional 4D5-8 single chain antibody with a (Gly4Ser3 linker precipitated at physiological pH 7.4. Conclusions This modular monomeric recombinant fluorescent antibody platform may be used to create a range of recombinant coloured antibody molecules for quantitative in situ, in vivo and ex vivo imaging, cell sorting and cell

  12. Expression of recombinant multi-coloured fluorescent antibodies in gor -/trxB - E. coli cytoplasm

    OpenAIRE

    Markiv Anatoliy; Beatson Richard; Burchell Joy; Durvasula Ravi V; Kang Angray S

    2011-01-01

    Abstract Background Antibody-fluorophore conjugates are invaluable reagents used in contemporary molecular cell biology for imaging, cell sorting and tracking intracellular events. However they suffer in some cases from batch to batch variation, partial loss of binding and susceptibility to photo-bleaching. In theory, these issues can all be addressed by using recombinant antibody fused directly to genetically encoded fluorescent reporters. However, single-chain fragment variable domains link...

  13. ANTIBODIES TO THE CYTOPLASM OF NEUTROPHILS: A MARKER OF UNFAVORABLE CLINICAL COURSE IN NON-SPECIFIC ULCERATIVE COLITIS

    Directory of Open Access Journals (Sweden)

    A. G. Kharitonov

    2010-01-01

    Full Text Available Antibodies to the cytoplasm of neutrophils (p-ANCA are detectable in 67% of patients with ulcerative colitis (UC. We have revealed typical clinical features of the patients with diagnostic p-ANCA titer, i.e., longer disease duration, prolonged fevers, and increased stool frequency. Moreover, thrombocytosis, leukocytosis and hypoalbuminemia are more common in this group. By endoscopic examination, mucosal ulcers of the colon are significantly more frequent in this group of patients. We have also noted higher rates of severe and relapsing cases among p-ANCA-positive patients. The data obtained allow us to suggest that the diagnostic titers of p-ANCA are predictive for unfavorable prognosis in UC.

  14. Anti-B cell antibody therapies for inflammatory rheumatic diseases

    DEFF Research Database (Denmark)

    Faurschou, Mikkel; Jayne, David R W

    2014-01-01

    Several monoclonal antibodies targeting B cells have been tested as therapeutics for inflammatory rheumatic diseases. We review important observations from randomized clinical trials regarding the efficacy and safety of anti-B cell antibody-based therapies for rheumatoid arthritis, systemic lupus...... erythematosus, antineutrophil cytoplasmic antibody-associated vasculitis, polymyositis/dermatomyositis, and primary Sjögren's syndrome. For some anti-B cell agents, clinical benefits have been convincingly demonstrated, while other B cell-targeted therapies failed to improve outcomes when added to standard...... and functions in rheumatic disorders. Future studies should also evaluate how to maintain disease control by means of conventional and/or biologic immunosuppressants after remission-induction with anti-B cell antibodies....

  15. Correlation of Antineutrophil Cytoplasmic Antibodies and Ulcerative Colitis%抗中性粒细胞胞质抗体与溃疡性结肠炎的关系

    Institute of Scientific and Technical Information of China (English)

    张玉; 徐三荣; 林庚金

    2000-01-01

    目的探讨血清抗中性粒细胞胞质抗体(ANCA)与溃疡性结肠炎的关系。方法应用间接免疫荧光法(IIF)及酶联免疫吸附试验(ELISA)测定了溃疡性结肠炎(UC)及非溃疡性结肠炎(NUC)患者的血清抗中性粒细胞胞质抗体(ANCA)。结果 ANCA阳性率分别为UC组10.53%,NUC组4.76%,两组相比无显著性差异。UC组ANCA阳性率与病情程度、病变范围、病程长短、病理严重程度均无明显关系(P>0.05)。结论我国UC患者ANCA阳性率不高的因素可能是:①被检患者人种不同,缺乏可比性;② UC病人中可能存在血清ANCA阴性亚群。

  16. Increased infectivity in human cells and resistance to antibody-mediated neutralization by truncation of the SIV gp41 cytoplasmic tail.

    Science.gov (United States)

    Kuwata, Takeo; Kaori, Takaki; Enomoto, Ikumi; Yoshimura, Kazuhisa; Matsushita, Shuzo

    2013-01-01

    The role of antibodies in protecting the host from human immunodeficiency virus type 1 (HIV-1) infection is of considerable interest, particularly because the RV144 trial results suggest that antibodies contribute to protection. Although infection of non-human primates with simian immunodeficiency virus (SIV) is commonly used as an animal model of HIV-1 infection, the viral epitopes that elicit potent and broad neutralizing antibodies to SIV have not been identified. We isolated a monoclonal antibody (MAb) B404 that potently and broadly neutralizes various SIV strains. B404 targets a conformational epitope comprising the V3 and V4 loops of Env that intensely exposed when Env binds CD4. B404-resistant variants were obtained by passaging viruses in the presence of increasing concentration of B404 in PM1/CCR5 cells. Genetic analysis revealed that the Q733stop mutation, which truncates the cytoplasmic tail of gp41, was the first major substitution in Env during passage. The maximal inhibition by B404 and other MAbs were significantly decreased against a recombinant virus with a gp41 truncation compared with the parental SIVmac316. This indicates that the gp41 truncation was associated with resistance to antibody-mediated neutralization. The infectivities of the recombinant virus with the gp41 truncation were 7,900-, 1,000-, and 140-fold higher than those of SIVmac316 in PM1, PM1/CCR5, and TZM-bl cells, respectively. Immunoblotting analysis revealed that the gp41 truncation enhanced the incorporation of Env into virions. The effect of the gp41 truncation on infectivity was not obvious in the HSC-F macaque cell line, although the resistance of viruses harboring the gp41 truncation to neutralization was maintained. These results suggest that viruses with a truncated gp41 cytoplasmic tail were selected by increased infectivity in human cells and by acquiring resistance to neutralizing antibody. PMID:23717307

  17. Increased infectivity in human cells and resistance to antibody-mediated neutralization by truncation of the SIV gp41 cytoplasmic tail

    Directory of Open Access Journals (Sweden)

    Takeo eKuwata

    2013-05-01

    Full Text Available The role of antibodies in protecting the host from human immunodeficiency virus type 1 (HIV-1 infection is of considerable interest, particularly because the RV144 trial results suggest that antibodies contribute to protection. Although infection of nonhuman primates with simian immunodeficiency virus (SIV is commonly used as an animal model of HIV-1 infection, the viral epitopes that elicit potent and broad neutralizing antibodies to SIV have not been identified. We isolated a monoclonal antibody (MAb B404 that potently and broadly neutralizes various SIV strains. B404 targets a conformational epitope comprising the V3 and V4 loops of Env that intensely exposed when Env binds CD4. B404-resistant variants were obtained by passaging viruses in the presence of increasing concentration of B404 in PM1/CCR5 cells. Genetic analysis revealed that the Q733stop mutation, which truncates the cytoplasmic tail of gp41, was the first major substitution in Env during passage. The maximal inhibition by B404 and other MAbs were significantly decreased against a recombinant virus with a gp41 truncation compared with the parental SIVmac316. This indicates that the gp41 truncation was associated with resistance to antibody-mediated neutralization. The infectivities of the recombinant virus with the gp41 truncation were 7900-fold, 1000-fold, and 140-fold higher than those of SIVmac316 in PM1, PM1/CCR5, and TZM-bl cells, respectively. Immunoblotting analysis revealed that the gp41 truncation enhanced the incorporation of Env into virions. The effect of the gp41 truncation on infectivity was not obvious in the HSC-F macaque cell line, although the resistance of viruses harboring the gp41 truncation to neutralization was maintained. These results suggest that viruses with a truncated gp41 cytoplasmic tail were selected by increased infectivity in human cells and by acquiring resistance to neutralizing antibody.

  18. Antibody-based analysis reveals “filamentous vs. non-filamentous” and “cytoplasmic vs. nuclear” crosstalk of cytoskeletal proteins

    Energy Technology Data Exchange (ETDEWEB)

    Kumeta, Masahiro, E-mail: kumeta@lif.kyoto-u.ac.jp [Graduate School of Biostudies, Kyoto University, Kyoto 606-8501 (Japan); Hirai, Yuya; Yoshimura, Shige H. [Graduate School of Biostudies, Kyoto University, Kyoto 606-8501 (Japan); Horigome, Tsuneyoshi [Graduate School of Science and Technology, Niigata University, Niigata 950-2181 (Japan); Takeyasu, Kunio [Graduate School of Biostudies, Kyoto University, Kyoto 606-8501 (Japan)

    2013-12-10

    To uncover the molecular composition and dynamics of the functional scaffold for the nucleus, three fractions of biochemically-stable nuclear protein complexes were extracted and used as immunogens to produce a variety of monoclonal antibodies. Many helix-based cytoskeletal proteins were identified as antigens, suggesting their dynamic contribution to nuclear architecture and function. Interestingly, sets of antibodies distinguished distinct subcellular localization of a single isoform of certain cytoskeletal proteins; distinct molecular forms of keratin and actinin were found in the nucleus. Their nuclear shuttling properties were verified by the apparent nuclear accumulations under inhibition of CRM1-dependent nuclear export. Nuclear keratins do not take an obvious filamentous structure, as was revealed by non-filamentous cytoplasmic keratin-specific monoclonal antibody. These results suggest the distinct roles of the helix-based cytoskeletal proteins in the nucleus. - Highlights: • A set of monoclonal antibodies were raised against nuclear scaffold proteins. • Helix-based cytoskeletal proteins were involved in nuclear scaffold. • Many cytoskeletal components shuttle into the nucleus in a CRM1-dependent manner. • Sets of antibodies distinguished distinct subcellular localization of a single isoform. • Nuclear keratin is soluble and does not form an obvious filamentous structure.

  19. T细胞受体α链稳定区基因-575A/G与原发抗中性粒细胞抗体相关性小血管炎的关联分析%Relationship of TCRCα-575A/G polymorphism with anti-neutrophil antibody associated vasculitis in Chinese Han population

    Institute of Scientific and Technical Information of China (English)

    薛超; 张益民; 廖蕴华; 杨桢华; 黄莉; 许佳; 项新

    2009-01-01

    Objective To investigate the relationship of TCRCα-575A/G polymorphism with anti-neutrophil antibody(ANCA) associated vasculitis in Chinese Han population. Methods 86 cases of ANCA associated vasculi-tis in Chinese Han population and 196 healthy subjects were enrolled. TCRCα-575A/G was genotyped by PCR-re-striction fragment length polymorphism (PCR-RFLP) assay. Case-control study was performed. Results No signifi-cant difference was found in either genotype distribution(AA,AG,GG) or allele frequencies between 86 patients and healthy subjects(P>0.05);But significant differences between AA group, AG group, and GG group in systolic pres-sure[(127.47±24.18)、(124.11±25.21)、(148.92±19.23) mm Hg],diastolic pressure [(75.35±14.12)、 (74.50±13.01)、(85.46±9.40) mm Hg],red blood cell count[(3.41±1.01)×109/L、(3.46±1.04)× 109/L、(2.68±0.67)×109/L] and hemoglobin [(90.45±20.69)、(100.66±29.80)、(77.61±15.81) g/L (P0.05 ). Conclusions In Chi-nese Han population,TCRCα-575A/G polymorphism might not be related to genetic susceptibility and chronic renal failure of ANCA associated vaseulitis;but G allele might be associated with more serious anaemia and hypertension.%目的 探讨T细胞受体α链稳定区(TCRCα)基因-575 A/G多态性与汉族人群原发抗中性粒细胞抗体(ANCA)相关性小血管炎的关联关系.方法 86例原发ANCA相关性小血管炎汉族患者与196例汉族健康人作对照,PCR-RFLP鉴定其TCRCα-575A/G基因型,进行病例-对照研究与临床分析.结果 86例患者中,TCRCα基因-575 A/G存在从、AG、GG 3种基因型和A、G 2种等位基因,与正常对照组相比,分布频率差异均无统计学意义(P均>0.05);TCRCα基因-575A/G基因型不同的患者组(AA组、AG组、GG组)相比,在收缩压[(127.47±24.18)、(124.11±25.21)、(148.92±19.23)mm Hg]、舒张压[(75.35±14.12)、(74.50±13.01)、(85.46±9.40)mm Hg]、红细胞数[(3.41±1.01)×109/L、(3.46±1.04)×109/L、(2.68±0.67)×109/L

  20. Cytoplasmic Z-RNA

    Energy Technology Data Exchange (ETDEWEB)

    Zarling, D.A.; Calhoun, C.J.; Hardin, C.C.; Zarling, A.H.

    1987-09-01

    Specific immunochemical probes for Z-RNA were generated and characterized to search for possible Z-RNA-like double helices in cells. Z-RNA was detected in the cytoplasm of fixed protozoan cells by immunofluorescence microscopy using these anti-Z-RNA IgCs. In contrast, autoimmune or experimentally elicited anti-DNA antibodies, specifically reactive with B-DNA or Z-DNA, stained the nuclei. Pre-or nonimmune IgGs did not bind to the cells. RNase A or T1 digestion eliminated anti-Z-RNA IgG binding to cytoplasmic determinants; however, DNase I or mung bean nuclease had no effect. Doxorubicin and ethidium bromide prevented anti-Z-RNA antibody binding; however, actinomycin D, which does not bind double-stranded RNA, did not. Anti-Z-RNA immunofluorescence was specifically blocked in competition assays by synthetic Z-RNA but not Z-DNA, A-RNA, or single-stranded RNAs. Thus, some cytoplasmic sequences in fixed cells exist in the left-handed Z-RNA conformation.

  1. Anti-hLAMP2-antibodies and dual positivity for anti-GBM and MPO-ANCA in a patient with relapsing pulmonary-renal syndrome

    Directory of Open Access Journals (Sweden)

    Kistler Thomas

    2011-06-01

    Full Text Available Abstract Background Pulmonary-renal syndrome associated with anti-glomerular basement membrane (GBM antibodies, also known as Goodpasture's syndrome, is a rare but acute and life-threatening condition. One third of patients presenting as anti-GBM antibody positive pulmonary-renal syndrome or rapidly progressive glomerulonephritis are also tested positive for anti-neutrophil cytoplasmic antibodies (ANCA. Whilst anti-GBM disease is considered a non-relapsing condition, the long-term course of double-positive patients is less predictable. Case Presentation We report a patient with such dual positivity, who presented with pulmonary hemorrhage, crescentic glomerulonephritis and membranous nephropathy. Plasmapheresis in combination with immunosuppresive therapy led to a rapid remission but the disease relapsed after two years. The serum of the patient was tested positive for antibodies to human lysosomal membrane protein 2 (hLAMP2, a novel autoantigen in patients with active small-vessel vasculitis (SVV. The anti-hLAMP2 antibody levels correlated positively with clinical disease activity in this patient. Conclusion We hypothesize that this antibody may indicate a clinical course similar to ANCA-associated vasculitis in double-positive patients. However, this needs to be confirmed on comprehensive patient cohorts.

  2. Prevalence and clinical significance of anti-lactoferrin autoantibodies in inflammatory bowel diseases and primary sclerosing cholangitis

    NARCIS (Netherlands)

    Roozendaal, C; Horst, G; Pogany, K; Kleibeuker, JH; Haagsma, EB; Limburg, PC; Kallenberg, CGM; Van Millegen de Wit, AWM; Spik, G; Legrand, D; Mazurier, J; Pierce, A; Perraudin, JP

    1998-01-01

    Anti-neutrophil cytoplasmic antibodies (ANCA) are autoantibodies directed against cytoplasmic constituents of neutrophil granulocytes. Antibodies with specificity for proteinase 3 and myeloperoxidase are seromarkers for systemic vasculitides. ANCA with specificity for lactoferrin were described in p

  3. Diagnosis of autoimmune neutropenia by neutrophil-bound IgG and IgM antibodies.

    Science.gov (United States)

    Ito, Taichi; Taniuchi, Shoichiro; Tsuji, Shoji; Iharada, Anna; Hasui, Masafumi; Kaneko, Kazunari

    2011-10-01

    Autoimmune neutropenia (AIN) in infancy is caused by antineutrophil (granulocyte-specific) autoantibodies. These antibodies are rarely found in circulation because their serum levels are extremely low. We hypothesized that a direct granulocyte immunofluorescence test (D-GIFT) that enables us to detect neutrophil-bound autoantibodies consisting of both immunoglobulin (Ig) G and IgM has better diagnostic value than the detection of circulating autoantibodies. Whole blood (100 μL) was obtained from 50 infants with AIN, 12 infants with transient neutropenia, and 37 control infants. D-GIFT was performed using both fluorescein isothiocyanate-conjugated antihuman IgG Fc portion monoclonal antibodies and fluorescein isothiocyanate antihuman IgM monoclonal antibodies. Results were assessed as relative fluorescence intensity (RFI). The RFIs of antineutrophil IgG-bound and antineutrophil IgM-bound cells in patients with AIN were significantly higher than those in patients with transient neutropenia and in controls. Positive results, as assessed by RFI scores of more than 1.81 in either antineutrophil IgG-bound or antineutrophil IgM-bound cells, showed the sensitivity and specificity of D-GIFT, and the areas under the receiver operating characteristic curve (0.98, 0.98, and 0.997, respectively) in the diagnosis of AIN. D-GIFT detecting both neutrophil-bound IgG autoantibodies and IgM autoantibodies has discriminatory power for identifying patients with AIN and, therefore, can be a useful diagnostic test. PMID:21941149

  4. Dysregulated neutrophil-endothelial interaction in antineutrophil cytoplasmic autoantibody (ANCA)-associated vasculitides : Implications for pathogenesis and disease intervention

    NARCIS (Netherlands)

    Hu, Nan; Westra, Johanna; Kallenberg, Cees G. M.

    2011-01-01

    The interplay between neutrophils and endothelial cells allows ANCA to become pathogenic and results in uncontrolled inflammation in the vessel wall. This review presents an overall view on neutrophil-endothelial interaction during inflammation with a focus on ANCA-associated vasculitis, and summari

  5. Cytoplasmic dynein nomenclature

    Science.gov (United States)

    Pfister, K. Kevin; Fisher, Elizabeth M.C.; Gibbons, Ian R.; Hays, Thomas S.; Holzbaur, Erika L.F.; McIntosh, J. Richard; Porter, Mary E.; Schroer, Trina A.; Vaughan, Kevin T.; Witman, George B.; King, Stephen M.; Vallee, Richard B.

    2005-01-01

    A variety of names has been used in the literature for the subunits of cytoplasmic dynein complexes. Thus, there is a strong need for a more definitive consensus statement on nomenclature. This is especially important for mammalian cytoplasmic dyneins, many subunits of which are encoded by multiple genes. We propose names for the mammalian cytoplasmic dynein subunit genes and proteins that reflect the phylogenetic relationships of the genes and the published studies clarifying the functions of the polypeptides. This nomenclature recognizes the two distinct cytoplasmic dynein complexes and has the flexibility to accommodate the discovery of new subunits and isoforms. PMID:16260502

  6. Anti-proteinase 3 antibodies in diffuse systemic sclerosis (SSc with normotensive renal impairment: is it suggestive for an overlapping between SSc and idiopathic vasculitis?

    Directory of Open Access Journals (Sweden)

    V. Campanella

    2011-09-01

    Full Text Available Objective. To test the prevalence of anti-neutrophil cytoplasmic antibodies (ANCA in systemic sclerosis (SSc and to verify a possible association of ANCA with normotensive renal involvement in SSc. Patients and methods: 51 patients affected by SSc, 35 with diffuse scleroderma (dSSc and 16 with limited scleroderma (lSSc, were tested for ANCA by indirect immunofluorescence (IIF on human ethanol and formalin-acetone-fixed granulocytes (before and after DNase treatment, by conventional enzyme linked immuno-sorbent assay (ELISA and by capture-ELISA. Results. Six out of 51 selected SSc patients had ANCA by IIF (11.7% and five presented a perinuclear/nuclear atypical ANCA pattern. In all cases we only found anti-proteinase3 (aPR3 antibodies. All ANCA positive patients had diffuse form of SSc (17.1%, all were anti-Scl70 positive (aScl70, five patients had proteinuria, three had microscopic haematuria. All ANCA positive patients were normotensive with normal renin plasma levels, the mean erythrocyte sedimentation rate (ESR was higher in this group compared to the other SSc patients. Conclusions. Our study shows that aPR3 is not rare in dSSc. According to the clinical and serological findings and to the recent literature, we can hypothesise that when ANCA are found in SSc, an overlapping of scleroderma with systemic necrotizing vasculitis should be suspected.

  7. Epitope analysis of anti-myeloperoxidase antibodies in patients with ANCA-associated vasculitis.

    Directory of Open Access Journals (Sweden)

    Shen-Ju Gou

    Full Text Available OBJECTIVE: Increasing evidences have suggested the pathogenic role of anti-neutrophil cytoplasmic antibodies (ANCA directing myeloperoxidase (MPO in ANCA-associated vasculitis (AAV. The current study aimed to analyze the association between the linear epitopes of MPO-ANCA and clinicopathological features of patients with AAV. METHODS: Six recombinant linear fragments, covering the whole length amino acid sequence of a single chain of MPO, were produced from E.coli. Sera from 77 patients with AAV were collected at presentation. 13 out of the 77 patients had co-existence of serum anti-GBM antibodies. Ten patients also had sequential sera during follow up. The epitope specificities were detected by enzyme-linked immunosorbent assay using the recombinant fragments as solid phase ligands. RESULTS: Sera from 45 of the 77 (58.4% patients with AAV showed a positive reaction to one or more linear fragments of the MPO chain. The Birmingham Vasculitis Activity Scores and the sera creatinine were significantly higher in patients with positive binding to the light chain fragment than that in patients without the binding. The epitopes recognized by MPO-ANCA from patients with co-existence of serum anti-GBM antibodies were mainly located in the N-terminus of the heavy chain. In 5 out of the 6 patients, whose sera in relapse recognize linear fragments, the reactivity to linear fragments in relapse was similar to that of initial onset. CONCLUSION: The epitope specificities of MPO-ANCA were associated with disease activity and some clinicopathological features in patients with ANCA-associated vasculitis.

  8. Absence of cross-reactivity to myeloperoxidase of anti-thyroid microsomal antibodies in patients with autoimmune thyroid diseases

    NARCIS (Netherlands)

    Freire, BA; Paula, ID; Paula, F; Kallenberg, GGM; Limburg, PC; Queluz, TT

    2001-01-01

    Background: Thyroperoxidase is the major antigen of the thyroid microsomal antibodies (TMA) detected in autoimmune thyroid diseases. Its amino acid sequence has 44% homology with myeloperoxidase (MPO), an enzyme present in the primary granules of neutrophils and one of the major antineutrophil cytop

  9. Cytoplasm Affects Embryonic Development

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    @@ Recent studies by CAS researchers furnish strong evidence that a fertilized egg's nucleus isn't the sole site of control for an embryo's development. A research team headed by Prof. Zhu Zuoyan from the CAS Institute of Hydrobiology in Wuhan discovered that cytoplasm affects the number of vertebrae in cloned offspring created when nuclei from one fish genus were transplanted to enucleated eggs of another.

  10. Cytoplasmic Male Sterility in Maize

    Institute of Scientific and Technical Information of China (English)

    RONG Ting-zhao; LI Wan-chen; CAO Mo-ju; HU Chang-yuan

    2002-01-01

    14 isoplasmic and allonuclear cytoplasmic male sterile lines were used as female parents, 8 tester lines as male parents, 101 F1 progenies were obtained. Fertility restoration response of 101 F1 progenies were investigated through field observation and pollen stainability examination under microscope. 14 isoplasmic and allonuclear cytoplasmic male sterile lines were developed by repeated backcross with recurrent male parent lines for more than 8 generations. The result shows: tester line Zifeng1 not only restored the isoplasmic and allonuclear sterile lines of group C backcrossed with Mo17, Yu30 and Heer, but also completely restored the isoplasmic and allonuclear cytoplasm male sterile lines of group T backcrossed with Mo17, HZS , 1792 ,292 and Yu30. Therefore, nuclear background limits the use of Zifeng1 as a tester for identification of cytoplasmic male sterility. Furthermore RFLPs of mitochondrial DNA of 6 isonuclear and alloplasmic cytoplasmic male sterile lines were analyzed with Bam H Ⅰ and Hind Ⅲ restriction endonuclease and mitochondrial DNA probes pBcmH3 and Cox Ⅱ. The same RFLPs were found within sterile cytoplasm of group C, including C,Chuan G, Lei 2 and Lei 3, but a different RFLP pattern was observed among sterile cytoplasm of group S, C,T and the normal cytoplasm. This result suggested that the RFLP markers tightly linked to sterile mitochondrial genes of different groups could be applied in the identifcation of cytoplasmic male sterility.

  11. Genetically distinct subsets within ANCA-associated vasculitis

    DEFF Research Database (Denmark)

    Lyons, Paul A; Rayner, Tim F; Trivedi, Sapna;

    2012-01-01

    Antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis is a severe condition encompassing two major syndromes: granulomatosis with polyangiitis (formerly known as Wegener's granulomatosis) and microscopic polyangiitis. Its cause is unknown, and there is debate about whether it is a sing...

  12. Characteristics and Outcomes of Granulomatosis With Polyangiitis (Wegener) and Microscopic Polyangiitis Requiring Renal Replacement Therapy: Results From the European Renal Association-European Dialysis and Transplant Association Registry

    NARCIS (Netherlands)

    Hruskova, Z.; Stel, V.S.; Jayne, D.; Aasarod, K.; Meester, J. de; Ekstrand, A.; Eller, K.; Heaf, J.G.; Hoitsma, A.J.; Jimenez, C. Martos; Ravani, P.; Wanner, C.; Tesar, V.; Jager, K.J.

    2015-01-01

    BACKGROUND: This study describes the incidence and outcomes of European patients requiring renal replacement therapy (RRT) for kidney failure due to antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). STUDY DESIGN: Cohort study. SETTING & PARTICIPANTS: 12 renal registries providi

  13. The role of myeloperoxidase in the pathogenesis of systemic vasculitis

    NARCIS (Netherlands)

    Rutgers, A; Heeringa, P; Tervaert, J W

    2004-01-01

    Wegener's granulomatosis, microscopic polyangiitis, Churg-Strauss syndrome and idiopathic pauci-immune necrotizing crescentic glomerulonephritis are strongly associated with the presence of anti-neutrophil cytoplasmic antibodies (ANCA). These ANCA-associated vasculitides can serologically be separat

  14. Brief Report

    DEFF Research Database (Denmark)

    Faurschou, Mikkel; Westman, Kerstin; Rasmussen, Niels;

    2012-01-01

    The NORAM (Nonrenal Wegener's Granulomatosis Treated Alternatively with Methotrexate [MTX]) trial demonstrated that MTX can replace cyclophosphamide (CYC) as remission-inducing treatment for patients with newly diagnosed early systemic antineutrophil cytoplasmic antibody-associated vasculitis. Du...

  15. The environment, geoepidemiology and ANCA-associated vasculitides

    NARCIS (Netherlands)

    Chen, Min; Kallenberg, Cees G. M.

    2010-01-01

    Anti-neutrophil cytoplasmic antibodies, directed against constituents of granules of neutrophils and lysosomes of monocytes, are serological markers of small vessel vasculitides, including Wegener's granulomatosis, microscopic polyangiitis, Churg-Strauss syndrome and renal-limited vasculitis. These

  16. Rituximab versus cyclophosphamide in ANCA-associated renal vasculitis : 2-year results of a randomised trial

    NARCIS (Netherlands)

    Jones, Rachel B.; Furuta, Shunsuke; Tervaert, Jan Willem Cohen; Hauser, Thomas; Luqmani, Raashid; Morgan, Matthew D.; Peh, Chen Au; Savage, Caroline O.; Segelmark, Marten; Tesar, Vladimir; van Paassen, Pieter; Walsh, Michael; Westman, Kerstin; Jayne, David R. W.; Stegeman, C. A.

    2015-01-01

    Objectives The RITUXVAS trial reported similar remission induction rates and safety between rituximab and cyclophosphamide based regimens for antineutrophil cytoplasm antibody (ANCA)-associated vasculitis at 12months; however, immunosuppression maintenance requirements and longer-term outcomes after

  17. Genetically Distinct Subsets within ANCA-Associated Vasculitis

    NARCIS (Netherlands)

    Lyons, Paul A.; Rayner, Tim F.; Trivedi, Sapna; Holle, Julia U.; Watts, Richard A.; Jayne, David R. W.; Baslund, Bo; Brenchley, Paul; Bruchfeld, Annette; Chaudhry, Afzal N.; Tervaert, Jan Willem Cohen; Deloukas, Panos; Feighery, Conleth; Gross, Wolfgang L.; Guillevin, Loic; Gunnarsson, Iva; Harper, Lorraine; Hruskova, Zdenka; Little, Mark A.; Martorana, Davide; Neumann, Thomas; Ohlsson, Sophie; Padmanabhan, Sandosh; Pusey, Charles D.; Salama, Alan D.; Sanders, Jan-Stephan F.; Savage, Caroline O.; Segelmark, Mrten; Stegeman, Coen A.; Tesar, Vladimir; Vaglio, Augusto; Wieczorek, Stefan; Wilde, Benjamin; Zwerina, Jochen; Rees, Andrew J.; Clayton, David G.; Smith, Kenneth G. C.

    2012-01-01

    BACKGROUND Antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis is a severe condition encompassing two major syndromes: granulomatosis with polyangiitis (formerly known as Wegener's granulomatosis) and microscopic polyangiitis. Its cause is unknown, and there is debate about whether it i

  18. Asbestos and Probable Microscopic Polyangiitis

    Directory of Open Access Journals (Sweden)

    George S Rashed Philteos

    2004-01-01

    Full Text Available Several inorganic dust lung diseases (pneumoconioses are associated with autoimmune diseases. Although autoimmune serological abnormalities are common in asbestosis, clinical autoimmune/collagen vascular diseases are not commonly reported. A case of pulmonary asbestosis complicated by perinuclear-antineutrophil cytoplasmic antibody (myeloperoxidase positive probable microscopic polyangiitis (glomerulonephritis, pericarditis, alveolitis, multineuritis multiplex is described and the possible immunological mechanisms whereby asbestosis fibres might be relevant in induction of antineutrophil cytoplasmic antibodies are reviewed in the present report.

  19. The clinical value of auto antibodies detection in diagnosis of primary sclerosing cholangitis%原发性硬化性胆管炎自身抗体检测的临床意义

    Institute of Scientific and Technical Information of China (English)

    谭立明; 彭卫华; 李华; 曹莉萍; 王园园; 熊金旺; 王外梅

    2011-01-01

    目的 探讨自身抗体检测对诊断原发性硬化性胆管炎(PSC)患者的临床意义.方法 采用间接免疫荧光法(IIF)检测抗核抗体(ANA)和抗中性粒细胞胞质抗体(ANCA)并对其结果进行回顾性分析.结果 PSC检测ANA、ANCA、c-ANCA和p-ANCA结果与对照组比较P<0.01,p-ANCA与c-ANCA组间比较P<0.01,差异有非常显著性意义.PSC患者与病毒性肝炎比较,除HCV以外,经x2检验,P<0.01差异均有非常显著性意义.结论 血清ANCA、p-ANCA抗体检测对诊断PSC有着重要作用.对提高PSC在临床上同其它肝病鉴别诊断亦非常重要.%To study the clinical value of the detections of auto antibodies in diagnosis of patients with primary sclerosing cholangitis (PSC). Anti -nuclear antibodies (ANA) and antineutrophil cytoplasmic antibodies (ANCA) were detected by IIF and the results were retrospectively analyzed. ANA, ANCA, c-ANCA and p-ANCA of PSC were higher than that of the controls (P < 0.01). And group comparison of p-ANCA and c-ANCA demonstrated significantly difference (P < 0.01). Cases with PSC and viral hepatitis, except hepatitis C, also showed significant difference of these antibodies (P < 0.01). Our results indicate that the method will be very useful for the diagnosis and differential diagnosis of PSC by detecting ANA and p-ANCA.

  20. The clinical significance of combined measurement of anti-saccharomyces cerevisia antibody and antiperinuclear neutrophil cytoplasmic antibody in inflammatory bowel disease%联合检测炎症性肠病患者抗酿酒酵母细胞抗体和抗中性粒细胞抗体的临床意义

    Institute of Scientific and Technical Information of China (English)

    张蜀澜; 李永哲; 李磊; 佟大伟; 胡朝军; 何纯; 李骥

    2008-01-01

    Objective To evaluate the clinical significance of combined measurement of antiperinuclear neutrophil cytoplasmic antibody(ANCA)and anti-saccharomyees cerevisia antibody(ASCA)for the diagnosis of inflammatory bowel disease(IBD)patients and difierentiation of Crohn's disease(CD)with ulcerative colitis(UC).Methods A total of 159 patients with IBD(97 UC,62 CD),167 patients with other non-IBD gastrointestinal conditions(NIBDC)and 25 healthy controls(HC)were recruited in our research.ASCA and ANCA were detected by enzyme-linked immunosorbent assay(ELISA)and indirect immunofluorescence assay.respectively.Results The prevalence of ASCA-IsA or IgG in CD group,UC group,NIBDC and HC were43.5%,14.4%,29.3%and 0,respectively.The prevalence of ASCA-IgA or IgG in CD group were higher than those in other groups(X2=16.76 or 4.12,P<0.01 or<0.05).The prevalence of ANCA in CD group.UC group,NIBDC and HC were 8.1%,56.7%,4.8%and 0,respectively.The prevalence of AMA in UC group were much higher than those in other groups(X2=38.08 or 90.47,P<0.01).The sensitivity specificity and positive predictive value(PPV)in ASCA+/ANCA-were 40.3%.93.8% and 80.6%,respectively,and in ANCA+/ASCA-were 48.5%,98.4% and 97.9%,respectively.Condusions ASCA or ANCA testing alone are not sensitive enoulgh for diagnosing CD and UC,but their combination asses are specific for differential diagnosis between CD and UC.Combined testing of ASCA-IgA with IgG can improve the sensitivity in screening CD patients.The ASCA positive pattern in Chinese CD group are correlated with surgery.%目的 探讨联合检测炎症性肠病(IBD)患者血清中抗酿酒酵母细胞抗体(ASCA)和抗中性粒细胞胞浆抗体(ANCA)对IBD诊断和鉴别诊断的应用价值.方法 用ELISA法和间接免疫荧光法分别测定159例IBD患者[溃疡性结肠炎(UC)97例,克罗恩病(CD)62例],167例主诉为腹痛、腹泻并除外IBD的患者和25名健康人血清中IgG型与IgA型ASCA和ANCA.结果 ASCA-IgA/IgG在CD组、UC组

  1. Cellular Subcompartments through Cytoplasmic Streaming.

    Science.gov (United States)

    Pieuchot, Laurent; Lai, Julian; Loh, Rachel Ann; Leong, Fong Yew; Chiam, Keng-Hwee; Stajich, Jason; Jedd, Gregory

    2015-08-24

    Cytoplasmic streaming occurs in diverse cell types, where it generally serves a transport function. Here, we examine streaming in multicellular fungal hyphae and identify an additional function wherein regimented streaming forms distinct cytoplasmic subcompartments. In the hypha, cytoplasm flows directionally from cell to cell through septal pores. Using live-cell imaging and computer simulations, we identify a flow pattern that produces vortices (eddies) on the upstream side of the septum. Nuclei can be immobilized in these microfluidic eddies, where they form multinucleate aggregates and accumulate foci of the HDA-2 histone deacetylase-associated factor, SPA-19. Pores experiencing flow degenerate in the absence of SPA-19, suggesting that eddy-trapped nuclei function to reinforce the septum. Together, our data show that eddies comprise a subcellular niche favoring nuclear differentiation and that subcompartments can be self-organized as a consequence of regimented cytoplasmic streaming.

  2. Photoreactivation of a Cytoplasmic Virus

    Science.gov (United States)

    Pfefferkorn, E. R.; Boyle, Mary K.

    1972-01-01

    Ultraviolet light-inactivated frog virus 3 is efficiently photoreactivated by chick embryo cells. A cellular enzyme is presumably responsible for this repair of viral deoxyribonucleic acid, for the phenomenon is insensitive to an inhibitor of protein synthesis and is not seen in mammalian cells that are known to lack photoreactivating enzyme. Since frog virus 3 is a cytoplasmic virus, functionally significant amounts of photoreactivating enzyme are probably present in the cytoplasm of chick embryo cells. PMID:5062749

  3. Analysis of the Detection Results of 1 318 Cases of Anti Neutrophil Cytoplasmic Antibodies%1318例抗中性粒细胞胞浆抗体检测结果分析

    Institute of Scientific and Technical Information of China (English)

    秦红群; 马稚强; 马顺高

    2016-01-01

    目的::探讨人群中抗中性粒细胞胞浆抗体(ANCA)的检出情况。方法:采用间接免疫荧光(IIF)法和 ELISA 法对1318例疑似原发性小血管炎患者做 ANCA、肝功能、肾功能指标检测,并进行分组统计分析。结果:1318例患者中检出 ANCA 阳性21例,阳性率为1.59%,其中 p-ANCA 阳性16例,阳性率1.21%,c-ANCA 阳性5例,阳性率0.38%;抗-MPO 阳性16例,阳性率1.21%;抗-PR3阳性5例,阳性率0.38%。对研究对象进行性别分组统计:男性患者635例,检出 ANCA 阳性11例占1.73%,女性患者683例,检出 ANCA 阳性10例占1.46%,两者无统计学差异(P >0.05);年龄分组统计:30岁以下184例,检出 ANCA 阳性1例,占0.54%,30岁以上1134例,检出 ANCA 阳性20例占1.76%,经χ2检验,两者有显著统计学差异(P 0.05);age group statistics:184 cases were under 30 years old,1 case of detection of ANCA positive(0.54%),above 30 years old in 1 134 cases detec-tion ANCA positive in 20 cases (1.76%),by χ2 test,both have a significant statistical difference (P <0.01 ),21 pa-tients with ANCA positive patients,1 7 cases of renal function was abnormal 80.95%.Conclusion:Local area suspected of primary small vessel vasculitis patients with ANCA detection rate was 1.59%,with P-ANCA and anti MPO mainly, positive rate of ANCA in the elderly group was significantly higher than that in children and adolescents group and ANCA antibodies and gender relations is not obvious;ANCA associated with most renal damage more serious.

  4. Anti-myeloperoxidase antibodies in patients with microscopic polyangiitis.

    Directory of Open Access Journals (Sweden)

    Pradhan V

    2003-11-01

    Full Text Available BACKGROUND & OBJECTIVES: Anti-neutrophil cytoplasmic antibodies in active necrotizing and crescentic glomerulonephritis are associated with systemic vasculitides like Wegener′s granulomatosis, Microscopic polyangitiis and Churg Strauss Syndrome. This study shows the incidence of ANCA with specificities to Myeloperoxidase and Proteinase3 in MPA cases and gives the correlation of ANCA with Birmingham Vasculitis Activity Score. MATERIAL & METHODS: Eighteen cases of MPA were diagnosed as per Chapel Hill Consensus Criteria. ANCA was detected by indirect immunofluorescence microscopy using fluorescence and Confocal Laser Scanning Microscopes. Anti-MPO and anti-PR3 were identified by commercial ELISAs and anti-MPO subclass and IgG isotypes were also detected. RESULTS: MPA patients showed a male preponderance with BVAS ranging from 17-30. Systemic involvement was seen in 88.9%, lower respiratory tract involvement in 77.8% and upper respiratory tract in only 33.3% cases. All these patients had perinuclear pattern on IIF, where titers ranged from 80-640 and ELISA showed anti-MPO; values ranging from 20-80 units/ml. IIF and ELISA showed a good correlation (r=0.77. Two patients having FPGN had dual specificities and had both anti-MPO and anti-PR3 which could be picked up only by ELISA. A good correlation (r=0.78 was observed between BVAS and ANCA levels as well. IgG ANCA was detected in 88.7% and 11.1% had IgG+IgM and IgG1+IgG4 ANCA was detected in 50% patients. CONCLUSION: p-ANCA with anti-MPO is highly specific for MPA; both IIF and ELISA should be carried out for true positivity and to identify rare cases of dual specificities. Confocal laser scanning microscopy is useful in identifying ANCA patterns especially when ANA is also positive. ANCA testing with BVAS assessment will surely help in early diagnosis and estimating the severity of this life threatening disease.

  5. Unusual extracardiac manifestations of isolated native tricuspid valve endocarditis

    OpenAIRE

    Wilczynska, M; Khoo, JP; McCann, GP

    2010-01-01

    Isolated native tricuspid valve endocarditis (TVE) in non-intravenous drug users is a very rare condition. We describe an unusual presentation of Enterococcus faecalis TVE associated with spondylodiscitis, positive cytoplasmic antineutrophil cytoplasmic antibodies and antiproteinase-3 antibodies vasculitic rash in an otherwise healthy patient with no history of intravenous drug use or underlying cardiac abnormalities.

  6. Detection of beta-tubulin in the cytoplasm of the interphasic Entamoeba histolytica trophozoites.

    Science.gov (United States)

    Gómez-Conde, Eduardo; Vargas-Mejía, Miguel Ángel; Díaz-Orea, María Alicia; Hernández-Rivas, Rosaura; Cárdenas-Perea, María Elena; Guerrero-González, Tayde; González-Barrios, Juan Antonio; Montiel-Jarquín, Álvaro José

    2016-08-01

    It is known that the microtubules (MT) of Entamoeba histolytica trophozoites form an intranuclear mitotic spindle. However, electron microscopy studies and the employment of anti-beta-tubulin (β-tubulin) antibodies have not exhibited these cytoskeletal structures in the cytoplasm of these parasites. The purpose of this work was to detect β-tubulin in the cytoplasm of interphasic E. histolytica trophozoites. Activated or non-activated HMI-IMSS-strain E. histolytica trophozoites were used and cultured for 72 h at 37 °C in TYI-S-33 medium, and then these were incubated with the anti-β-tubulin antibody of E. histolytica. The anti-β-tubulin antibody reacted with the intranuclear mitotic spindle of E. histolytica-activated trophozoites as control. In contrast, in non-activated interphasic parasites, anti-β-tubulin antibody reacted with diverse puntiform structures in the cytoplasm and with ring-shaped structures localized in the cytoplasm, cellular membrane and endocytic stomas. In this work, for the first time, the presence of β-tubulin is shown in the cytoplasm of E. histolytica trophozoites. PMID:27156446

  7. Thyroid Antibodies

    Science.gov (United States)

    ... be limited. Home Visit Global Sites Search Help? Thyroid Antibodies Share this page: Was this page helpful? Also known as: Thyroid Autoantibodies; Antithyroid Antibodies; Antimicrosomal Antibody; Thyroid Microsomal Antibody; ...

  8. Specificities of anti-neutrophil autoantibodies in patients with rheumatoid arthritis (RA)

    DEFF Research Database (Denmark)

    Brimnes, J; Halberg, P; Jacobsen, Søren;

    1997-01-01

    The objective of this study was to characterize antigens recognized by neutrophil-specific autoantibodies from patients with RA. Sera from 62 RA patients were screened by indirect immunofluorescence (IIF). Positive sera were further tested by ELISAs for antibodies against various granule proteins...

  9. [Antinuclear antibodies].

    Science.gov (United States)

    Cabiedes, Javier; Núñez-Álvarez, Carlos A

    2010-01-01

    Anti-nuclear antibodies (ANA) are immunoglobulin directed against autologous cell nuclear and cytoplasmic components. Besides the autoimmune ANA there are other ANA that can be detected in circulation, like natural and infectious ANA. Because of its high sensibility, detection of the ANA must be done by indirect immunofluorescence (IIF) as screening test and all of those positive samples are convenient to confirm its specificity by ELISA, western blot or other techniques. Positive ANA detected by IIF must be evaluated taking in to account the pattern and titer. The following recommended step is the specificity characterization (reactivity against extractable nuclear antigens [ENA], dsDNA, etc.) which is useful for the diagnosis and follow up of patients with autoimmune diseases, and by such reasoning, its detection must be performed in an orderly and reasonable way using guides or strategies focused to the good use and interpretation of the autoantibodies. The objective of this review is to present a compilation of the literature and our experience in the detection and study of the ANA.

  10. Detection of cytoplasmic proteins from Helicobacter pylori in Colony Lift Immunoassay.

    Science.gov (United States)

    Rojas-Rengifo, Diana F; Jaramillo, Carlos A; Haas, Rainer; Jiménez-Soto, Luisa F

    2015-12-01

    Use of the Colony Lift Immunoassay has been described for several Gram negative bacteria of medical interest. In all cases detection was limited to the use of antibodies against outer membrane proteins. Here we describe the adaptation of this method for detection of the cytoplasmic CagA toxin from Helicobacter pylori.

  11. Regulation of actin-dependent cytoplasmic motility by type II phytochrome occurs within seconds in Vallisneria gigantea epidermal cells.

    Science.gov (United States)

    Takagi, Shingo; Kong, Sam-Geun; Mineyuki, Yoshinobu; Furuya, Masaki

    2003-02-01

    The effects of light on actin-dependent cytoplasmic motility in epidermal cells of green leaves of the aquatic angiosperm Vallisneria gigantea were investigated quantitatively using a custom-made dynamic image analyzer. Cytoplasmic motility was measured by monitoring changes in the brightness of individual pixels on digitized images taken sequentially under infrared light. Acceleration and deceleration of cytoplasmic motility were regulated photoreversibly by type II phytochrome(s). This phytochrome-dependent induction of cytoplasmic motility did not occur uniformly in cytoplasm but took place as scattered patches in which no particular organelles, including nucleus, existed. The induction became detectable at 2.5 s after the start of irradiation with pulsed red light. In cells exposed to microbeam irradiation, cytoplasmic motility was induced only in sites in the cytoplasm that were irradiated directly, whereas nonirradiated neighboring areas were unaffected. The effect was short-lived, disappearing within a few minutes, and no signal was transmitted from an irradiated cell to its neighbors. Anti-phytochrome antibody-responsive protein(s) was detectable in the leaf extract by immunoblot and zinc blot analyses and in cryosections of the epidermis by immunocytochemistry. Although the phytochrome-dependent cytoplasmic motility was blocked by exogenously applied latrunculin B or cytochalasins, treatment of the dark-adapted cells with Ca(2+)-chelating reagents induced the cytoplasmic motility. We have proposed a model for the phytochrome regulation of cytoplasmic motility as one of the earliest responses to a light stimulus. PMID:12566576

  12. An AFM Observation on Fossil Cytoplasm

    Institute of Scientific and Technical Information of China (English)

    WANG Xin; YU Junping; FANG Xiaohong

    2008-01-01

    Fossil cytoplasm is a new research topic of interest in paleobotany. Atomic force microscope (AFM) is a new technology applied widely in physics and biology; however, it is rarely used in paleontology. Here we applied AFM for the first time to study fossil cytoplasm. The results indicate that the fossil cytoplasm is heterogeneous and full of ultrastructures, just like extant cytoplasm, and that the application of AFM, especially in combination with other techniques, can reveal the subcellular details of fossil plants with more confidence.

  13. Cytoplasmic Streaming - Skylab Student Experiment ED-63

    Science.gov (United States)

    1973-01-01

    This chart describes the Skylab student experiment (ED-63), Cytoplasmic Streaming, proposed by Cheryl A. Peitz of Arapahoe High School, Littleton, Colorado. Experiment ED-63 was to observe the effect of zero-gravity on cytoplasmic streaming in the aquatic plant named Elodea, commonly called water weed or water thyme. The phenomenon of cytoplasmic streaming is not well understood, but it is recognized as the circulation mechanism of the internal materials or cytoplasm of a cell. Cytoplasm is a gelatinous substance that has the ability to change its viscosity and flow, carrying various cell materials with it. The activity can be stimulated by sunlight or heat. In March 1972, NASA and the National Science Teachers Association selected 25 experiment proposals for flight on Skylab. Science advisors from the Marshall Space Flight Center aided and assisted the students in developing the proposals for flight on Skylab.

  14. 抗c-mpl抗体在系统性红斑狼疮并发血小板减少中的作用%Role of anti c-mpl antibody in systemic lupus erythematosus with thrombocytopenia

    Institute of Scientific and Technical Information of China (English)

    杨拓; 黄慈波; 赖蓓; 赵丽珂; 陈颖娟; 赵籥陶; 张春媚; 曾小峰

    2012-01-01

    目的:分析抗血小板生成素( thrombopoietin,TPO)受体(c-mpl)抗体在系统性红斑狼疮(systemic lupus erythematosus,SLE)血小板减少中的作用,探讨该抗体的致病机制及临床价值.方法:应用间接ELISA法检测24例伴血小板减少SLE患者(A组)、27例曾有血小板减少现已恢复正常SLE患者(B组)、18例无血小板减少SLE患者(C组)及18例正常对照者血清中抗c-mpl抗体、TPO水平,分析其与临床表现及SLE疾病活动性指数(SLE disease activity index,SLEDAI)之间的关系.结果:血清抗c-mpl抗体在SLE总的阳性率为18.8%,抗c-mpl抗体在A组、B组和C组中的阳性率差异无统计学意义(P=0.600).抗c-mpl抗体阳性组血小板计数明显低于阴性组(P=0.025),血清TPO水平明显高于阴性组(P =0.038),抗c-mpl抗体阳性与阴性组补体C3和C4、红细胞沉降率(erytrocyte sedimentation rate,ESR)、C反应蛋白(C-reactive protein,CRP)及抗核抗体(anti-nuclear antibody,ANA)、抗双链DNA抗体(anti double-strained DNA,dsDNA)、抗中性粒细胞胞浆抗体(antineutrophil cytoplasmic antibody,ANCA)阳性率、SLEDAI评分差异均无统计学意义(P>0.05).结论:抗c-mpl抗体可能通过与TPO竞争结合c-mpl,阻断TPO-c-mpl通路,引起巨核细胞分化、发育、成熟障碍,从而引起血小板减少.%Objective;To determine whether anti-thrompoietin receptor (TPO-R, c-mpl) antibody contributes to thrombocytopenia in systemic lupus erytematosus (SLE) and explore the pathogenic role of this antibody. Methods; Sera from 24 SLE patients with thrombocytopenia, 27 SLE patients having normal platelet counts with a history of thrombocytopenia, 18 SLE patients with neither thrombocytopenia nor post thrombocytopenia and 18 healthy controls were collected. Anti c-mpl antibodies were detected by an indi-rected ELISA assay. The serum TPO levels were measured by an ELISA assay. Clinical findings, au-toantibody profiles, and SLEDAI were evaluated. Results; Serum anti c

  15. Xenopus egg cytoplasm with intact actin.

    Science.gov (United States)

    Field, Christine M; Nguyen, Phuong A; Ishihara, Keisuke; Groen, Aaron C; Mitchison, Timothy J

    2014-01-01

    We report optimized methods for preparing Xenopus egg extracts without cytochalasin D, that we term "actin-intact egg extract." These are undiluted egg cytoplasm that contains abundant organelles, and glycogen which supplies energy, and represents the least perturbed cell-free cytoplasm preparation we know of. We used this system to probe cell cycle regulation of actin and myosin-II dynamics (Field et al., 2011), and to reconstitute the large, interphase asters that organize early Xenopus embryos (Mitchison et al., 2012; Wühr, Tan, Parker, Detrich, & Mitchison, 2010). Actin-intact Xenopus egg extracts are useful for analysis of actin dynamics, and interaction of actin with other cytoplasmic systems, in a cell-free system that closely mimics egg physiology, and more generally for probing the biochemistry and biophysics of the egg, zygote, and early embryo. Detailed protocols are provided along with assays used to check cell cycle state and tips for handling and storing undiluted egg extracts.

  16. A Challenging Twist in Pulmonary Renal Syndrome

    Directory of Open Access Journals (Sweden)

    Rajaie Namas

    2014-01-01

    Full Text Available Case. We report a rare case of hydralazine-induced anti-neutrophil cytoplasmic antibody-associated vasculitis. A 75-year-old African American woman with history of high blood pressure on hydralazine for 3 years presented with acute onset of shortness of breath and hemoptysis. Lab workup revealed a severe normocytic anemia and a serum creatinine of 5.09 mg/dL (baseline 0.9. Bronchoscopy demonstrated active pulmonary hemorrhage. A urine sample revealed red cell casts and a renal biopsy demonstrated pauci-immune, focally necrotizing glomerulonephritis with small crescents consistent with possible anti-neutrophil cytoplasmic antibody-positive renal vasculitis. Serologies showed high-titer MPO-ANCA and high-titer anti-histone antibodies. She was treated with intravenous steroids and subsequently with immunosuppression after cessation of hydralazine. The patient was subsequently discharged from hospital after a rapid clinical improvement. Conclusion. Hydralazine-induced anti-neutrophil cytoplasmic antibody-positive renal vasculitis is a rare adverse effect and can present as a severe vasculitic syndrome with multiple organ involvement. Features of this association include the presence of high titer of anti-myeloperoxidase anti-neutrophil cytoplasmic antibody with multiantigenicity, positive anti-histone antibodies, and the lack of immunoglobulin and complement deposition. Prompt cessation of hydralazine may be sufficient to reverse disease activity but immunosuppression may be needed.

  17. Detection of cytoplasmic glycosylation associated with hydroxyproline.

    Science.gov (United States)

    West, Christopher M; van der Wel, Hanke; Blader, Ira J

    2006-01-01

    A special class of glycosylation occurs on a proline residue of the cytoplasmic/nuclear protein Skp1 in the social amoeba Dictyostelium. For this glycosylation to occur, the proline must first be hydroxylated by the action of a soluble prolyl 4-hydroxylase acting on the protein. Cytoplasmic prolyl 4-hydroxylases are dioxygen-dependent enzymes that have low affinity for their O2 substrate and, therefore, have been implicated in O2-sensing in Dictyostelium, as well as in vertebrates and invertebrates. The sugar-hydroxyproline linkage has low abundance, is resistant to alkali cleavage and known glycosidases, and does not bind known lectins. However, initial screens for this modification can be made by assessing changes in electrophoretic mobility of candidate proteins after treatment of cells with prolyl hydroxylase inhibitors, and/or by metabolic labeling with [3H]sugar precursors. In addition, cytoplasmic hydroxylation/glycosylation can be assessed by assaying for cytoplasmic glycosyltransferases. Here we describe these methods and examples of their use in analyzing Skp1 glycosylation in Dictyostelium and the apicomplexan Toxoplasma gondii, the causative agent of toxoplasmosis in humans. PMID:17132515

  18. How crowded is the prokaryotic cytoplasm?

    NARCIS (Netherlands)

    Spitzer, Jan; Poolman, Bert; Ferguson, Stuart

    2013-01-01

    We consider biomacromolecular crowding within the cytoplasm of prokaryotic cells as a two-phase system of 'supercrowded' cytogel and 'dilute' cytosol; we simplify and quantify this model for a coccoid cell over a wide range of biomacromolecular crowding. The key result shows that the supercrowded cy

  19. Subunit organization in cytoplasmic dynein subcomplexes

    Science.gov (United States)

    King, Stephen J.; Bonilla, Myriam; Rodgers, Michael E.; Schroer, Trina A.

    2002-01-01

    Because cytoplasmic dynein plays numerous critical roles in eukaryotic cells, determining the subunit composition and the organization and functions of the subunits within dynein are important goals. This has been difficult partly because of accessory polypeptide heterogeneity of dynein populations. The motor domain containing heavy chains of cytoplasmic dynein are associated with multiple intermediate, light intermediate, and light chain accessory polypeptides. We examined the organization of these subunits within cytoplasmic dynein by separating the molecule into two distinct subcomplexes. These subcomplexes were competent to reassemble into a molecule with dynein-like properties. One subcomplex was composed of the dynein heavy and light intermediate chains whereas the other subcomplex was composed of the intermediate and light chains. The intermediate and light chain subcomplex could be further separated into two pools, only one of which contained dynein light chains. The two pools had distinct intermediate chain compositions, suggesting that intermediate chain isoforms have different light chain–binding properties. When the two intermediate chain pools were characterized by analytical velocity sedimentation, at least four molecular components were seen: intermediate chain monomers, intermediate chain dimers, intermediate chain monomers with bound light chains, and a mixture of intermediate chain dimers with assorted bound light chains. These data provide new insights into the compositional heterogeneity and assembly of the cytoplasmic dynein complex and suggest that individual dynein molecules have distinct molecular compositions in vivo. PMID:11967380

  20. The primary structure of rat brain (cytoplasmic) dynein heavy chain, a cytoplasmic motor enzyme.

    OpenAIRE

    Zhang, Z.; Tanaka, Y.; Nonaka, S; Aizawa, H.; Kawasaki, H; Nakata, T; Hirokawa, N

    1993-01-01

    Overlapping cDNA clones encoding the heavy chain of rat brain cytoplasmic dynein have been isolated. The isolated cDNA clones contain an open reading frame of 13,932 bp encoding 4644 aa (M(r), 532,213). The deduced protein sequence of the heavy chain of rat brain dynein shows significant similarity to sea urchin flagellar beta-dynein (27.0% identical) and to Dictyostelium cytoplasmic dynein (53.5% identical) throughout the entire sequence. The heavy chain of rat brain (cytoplasmic) dynein con...

  1. Cytoplasmic inheritance in Nicotiana tabacum L. autoradiography

    International Nuclear Information System (INIS)

    Tobacco pollen grains were labelled with leucine 14C, part then being used for artificial pollination of specific Nicotiana lines, while the rest were processed directly, as controls, for light and electron microscopy and autoradiography. The apparent differences in silver grain density observed in the controls may be related to the different physiological states of pollen grains during labelling. A radioactive precursor does not impair the viability and fertility of pollen grains. Samples taken and observed at successive intervals after pollination allowed the monitoring of cytoplasmic transmission to the embryo sac. The cytoplasmic materials remain confined within the micropylar canal and the embryo sac for several days after pollination. The tagged mitochondria found in the embryo sac are not necessarily of male origin, since the radioactive materials they contain might be products of metabolism. (author)

  2. Disassembly of simian virus 40 during passage through the endoplasmic reticulum and in the cytoplasm.

    Science.gov (United States)

    Kuksin, Dmitry; Norkin, Leonard C

    2012-02-01

    The nonenveloped polyomavirus simian virus 40 (SV40) is taken up into cells by a caveola-mediated endocytic process that delivers the virus to the endoplasmic reticulum (ER). Within the ER lumen, the capsid undergoes partial disassembly, which exposes its internal capsid proteins VP2 and VP3 to immunostaining with antibodies. We demonstrate here that the SV40 genome does not become accessible to detection while the virus is in the ER. Instead, the genome becomes accessible two distinct detection procedures, one using anti-bromodeoxyuridine antibodies and the other using a 5-ethynyl-2-deoxyuridine-based chemical reaction, only after the emergence of partially disassembled SV40 particles in the cytoplasm. These cytoplasmic particles retain some of the SV40 capsid proteins, VP1, VP2, and VP3, in addition to the viral genome. Thus, SV40 particles undergo discrete disassembly steps during entry that are separated temporally and topologically. First, a partial disassembly of the particles occurs in the ER, which exposes internal capsid proteins VP2 and VP3. Then, in the cytoplasm, disassembly progresses further to also make the genomic DNA accessible to immune detection.

  3. Evaluation of the Ca 1 antibody in the diagnosis of invasive breast cancer.

    OpenAIRE

    Clough, D G; Coghill, G R; Holley, M. P.

    1984-01-01

    An evaluation of Ca 1 antibody staining was performed on paraffin sections from 136 breast lesions (64 benign and 72 malignant). Although cytoplasmic staining was encountered significantly more often in malignant lesions, the false negative rate was 6.9% and the false positive rate 56.2%. Benign lesions which showed positive staining included gynaecomastia, cystic mastopathy and fibroadenomata. Various other monoclonal antibodies showed staining similar to Ca 1 antibody. Ca 1 antibody was obs...

  4. The maturation of antibody technology for the HIV epidemic.

    Science.gov (United States)

    Winnall, Wendy R; Beasley, Matthew D; Center, Rob J; Parsons, Matthew S; Kiefel, Ben R; Kent, Stephen J

    2014-08-01

    Antibodies are one of our most useful biological tools. Indeed, improvements in antibody-based technologies have ushered in a new era of antibody-based therapeutics, research and diagnostic tools. Although improved technologies have led to the development of therapeutic antibodies for treatment of malignancies and inflammatory conditions, the use of advanced antibody technology in the therapy of viral infections is in its infancy. Non-human primate studies have demonstrated that antibodies against the HIV envelope can both prevent viral infection and control viremia. Despite the obvious potential of antibody therapies against HIV, there remain limitations in production and purification capacity that require further research. Recent advances in recombinant antibody technology have led to the development of a range of novel antibody fragments, such as single-domain nanobodies and bispecific antibodies, that are capable of targeting cancer cells to cytotoxic T cells. Novel antibody production techniques have also been designed, allowing antibodies to be obtained from non-mammalian cells, bovine colostrum and the periplasm and cytoplasm of bacteria. These advances may allow large-scale production of HIV antibodies that are capable of protecting against HIV infection or serving as therapeutics that reduce the need for life-long antiretroviral treatment. This review summarises recent advances in antibody-based technologies and discusses the possibilities and challenges of using these advances to design prophylactics and therapeutics against HIV. PMID:24797582

  5. The Composition and Organization of Cytoplasm in Prebiotic Cells

    OpenAIRE

    Trevors, Jack T.

    2011-01-01

    This article discusses the hypothesized composition and organization of cytoplasm in prebiotic cells from a theoretical perspective and also based upon what is currently known about bacterial cytoplasm. It is unknown if the first prebiotic, microscopic scale, cytoplasm was initially contained within a primitive, continuous, semipermeable membrane, or was an uncontained gel substance, that later became enclosed by a continuous membrane. Another possibility is that the first cytoplasm in prebio...

  6. An atypical case of Wegener's granulomatosis complicated by sepsis and coxitis

    International Nuclear Information System (INIS)

    There is presented an atypical case Wegener's granulomatosis of maxillo-nasal region without bony destruction. The case is unusual because of sepsis and purulent coxitis. The patient responded well to treatment with vincristine and cyclophosphamide. ANCA (anti-neutrophil cytoplasmic antibodies) assays have very good sensitivity and specificity for Wegener's granulomatosis. (author)

  7. Interference of PR3-ANCA with the enzymatic activity of PR3 : differences in patients during active disease or remission of Wegener's granulomatosis

    NARCIS (Netherlands)

    Van der Geld, YM; Tool, ATJ; Videler, J.; De Haas, M; Tervaert, JWC; Stegeman, CA; Limburg, PC; Kallenberg, CGM; Roos, D

    2002-01-01

    Anti-neutrophil cytoplasmic antibodies (ANCA) against proteinase 3 (PR3) are strongly associated with Wegener's granulomatosis (WG) and are thought to be involved in its pathogenesis. Levels of PR3-ANCA do not always correspond to clinical disease activity. To investigate the relationship between fu

  8. Prognosis of microscopic polyangiitis with renal involvement: report of 60 Chinese patients

    Institute of Scientific and Technical Information of China (English)

    HU Wei-xin; LIU Zhi-hong; LIU Chun-bei; TANG Zheng; WANG Qing-wen; CHEN Hui-ping; LI Lei-shi

    2005-01-01

    @@ With the widespread clinical application of renal biopsy and serum antineutrophil cytoplasmic antibody (ANCA) testing, the percentage of renal injuries caused by systemic small vessel vasculitis (SVV) including microscopic polyangiitis (MPA)and Wegener's granulomatosis (WG) is on the rise in China.

  9. Connective tissue growth factor and the cicatrization of cellular crescents in ANCA-associated glomerulonephritis

    NARCIS (Netherlands)

    Hilhorst, Marc; Kok, Helena M.; Broekhuizen, Roel; van Paassen, Pieter; van Breda Vriesman, Peter; Goldschmeding, Roel; Nguyen, Tri Q; Cohen Tervaert, Jan Willem

    2015-01-01

    BACKGROUND: Outcome in patients with anti-neutrophil cytoplasmic antibodies (ANCA)-associated glomerulonephritis (AGN) is difficult to predict. Scoring of renal biopsies has significant but limited predictive value. We investigated whether analysis of plasma and urine levels, and immunostaining of b

  10. ANCA-associated glomerulonephritis : insights into etiology, pathogenesis, and prognosis

    NARCIS (Netherlands)

    Lind van Wijngaarden, Robert Arie Frederick de

    2009-01-01

    The body against itself When the immune system derails, antibodies directed against patients’ own molecules can be formed. This happens in ANCA-associated vasculitis, a vascular inflammation where antineutrophil cytoplasm autoantibodies (ANCA) cause damage, predominantly in the kidney. Untreated, va

  11. Refractory vasculitis

    NARCIS (Netherlands)

    Rutgers, Bram; Kallenberg, Cees G. M.

    2011-01-01

    Refractory vasculitis occurs in 4-5% of patients with anti-neutrophil cytoplasmic antibody associated vasculitis (AAV). Differences between therapies used for refractory disease are mostly reflected in the percentages of complete and partial remissions, but also in the number of serious side effects

  12. Mechanisms of vasculitis : How pauci-immune is ANCA-associated renal vasculitis?

    NARCIS (Netherlands)

    van Paassen, P.; Tervaert, J. W. Cohen; Heeringa, P.

    2007-01-01

    Both the innate and the acquired immune system are involved in the pathophysiology of renal vasculitis. However, anti-neutrophil cytoplasmic antibody (ANCA)-associated renal vasculitis is characterized by a 'pauci-immune' pattern of immunofluorescence during kidney biopsy, indicating the relative la

  13. Asbestos and Probable Microscopic Polyangiitis

    OpenAIRE

    George S Rashed Philteos; Kelly Coverett; Rajni Chibbar; Ward, Heather A; Cockcroft, Donald W

    2004-01-01

    Several inorganic dust lung diseases (pneumoconioses) are associated with autoimmune diseases. Although autoimmune serological abnormalities are common in asbestosis, clinical autoimmune/collagen vascular diseases are not commonly reported. A case of pulmonary asbestosis complicated by perinuclear-antineutrophil cytoplasmic antibody (myeloperoxidase) positive probable microscopic polyangiitis (glomerulonephritis, pericarditis, alveolitis, multineuritis multiplex) is described and the possible...

  14. Detection of immune deposits in skin lesions of patients with Wegener's granulomatosis

    NARCIS (Netherlands)

    Brons, RH; de Jong, MCJM; de Boer, NK; Stegeman, CA; Kallenberg, CGM; Tervaert, JWC

    2001-01-01

    Background-Wegener's granulomatosis (WG) is considered a pauci-immune systemic vasculitis based on the absence of immune deposits in renal biopsies of patients with active disease. In animal models of antineutrophil cytoplasmic antibody (ANCA) associated glomerulonephritis, immune deposits along the

  15. Genetically distinct subsets within ANCA-associated vasculitis.

    LENUS (Irish Health Repository)

    Lyons, Paul A

    2012-07-19

    Antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis is a severe condition encompassing two major syndromes: granulomatosis with polyangiitis (formerly known as Wegener\\'s granulomatosis) and microscopic polyangiitis. Its cause is unknown, and there is debate about whether it is a single disease entity and what role ANCA plays in its pathogenesis. We investigated its genetic basis.

  16. An atypical case of Wegener`s granulomatosis complicated by sepsis and coxitis; Ziarniniakowatosc Wegenera o nietypowej lokalizacji powiklana posocznica i ropnym zapaleniem stawu biodrowego

    Energy Technology Data Exchange (ETDEWEB)

    Makowski, A.; Faflik, J. [Centrum Zdrowia Matki Polki, Lodz (Poland)

    1993-12-31

    There is presented an atypical case Wegener`s granulomatosis of maxillo-nasal region without bony destruction. The case is unusual because of sepsis and purulent coxitis. The patient responded well to treatment with vincristine and cyclophosphamide. ANCA (anti-neutrophil cytoplasmic antibodies) assays have very good sensitivity and specificity for Wegener`s granulomatosis. (author). 17 refs, 2 figs.

  17. Re-evaluation of the histopathologic classification of ANCA-associated glomerulonephritis : a study of 121 patients in a single center

    NARCIS (Netherlands)

    Chang, Dong-yuan; Wu, Li-hua; Liu, Gang; Chen, Min; Kallenberg, Cees G. M.; Zhao, Ming-Hui

    2012-01-01

    Background. The recently published histopathologic classification of antineutrophil cytoplasmic antibody (ANCA)associated glomerulonephritis might greatly aid in the prognostication of patients at the time of diagnosis. This study aims to re-evaluate the new classification for its prognostic capacit

  18. Androgen deficiency in male patients diagnosed with ANCA-associated vasculitis : A cause of fatigue and reduced health-related quality of life?

    NARCIS (Netherlands)

    Tuin, Janneke; Sanders, Jan-Stephan F.; Buhl, Birgit M.; van Beek, Andre P.; Stegeman, Coen A.

    2013-01-01

    Introduction: Low testosterone levels in men are associated with fatigue, limited physical performance and reduced health-related quality of life (HRQOL); however, this relationship has never been assessed in patients with anti-neutrophil cytoplasmic antibodies (ANCA) -associated vasculitides (AAV).

  19. Hydralazine-induced constrictive pericarditis

    NARCIS (Netherlands)

    Franssen, CFC; ElGamal, MIH; Gans, ROB; Hoorntje, SJ

    1996-01-01

    A 59-year-old man was diagnosed as having constrictive pericarditis 17 months after a typical hydralazine-induced autoimmune syndrome, This late complication of hydralazine has been reported only once. Ten years later the patient was found to have anti-neutrophil cytoplasmic antibodies directed agai

  20. Molecular classification of Maize cytoplasms in a breeding program

    Directory of Open Access Journals (Sweden)

    Colombo. N * , Presello, D.A. , Kandus M. , G.E. Eyherabide and J.C. Salerno

    2012-06-01

    Full Text Available Cytoplasmic male sterility (CMS is maternally inherited in most of higher plants species. Together with nuclear restorer genes (Rf, CMS cytoplasms contribute significantly to the efficient production of hybrid seed. Three main types of male sterile cytoplasms are known in maize: T, S and C, which can be distinguished by crossing with specific restorer lines. Recently, PCR markers have been developed allowing the identification of different cytoplasms quickly and accurately. Our objective was to classify the cytoplasm type of maize inbred lines used in our breeding program and F1s obtained from crosses between CMS lines and elite maize lines using PCR multiplex. A multiplex PCR protocol was optimized for our conditions. We obtained the molecular classification of the analyzed cytoplasms. The optimized protocol is a valuable tool to trace male sterile cytoplasms and determine hybrid seed purity in our maize breeding program.

  1. Review article : The role of CD4(+) T cells in ANCA-associated systemic vasculitis

    NARCIS (Netherlands)

    Abdulahad, Wayel H.; Stegeman, Coen A.; Kallenberg, Cees G. M.

    2009-01-01

    Antineutrophil cytoplasmic autoantibody (ANCA)-associated systemic vasculitis (AASV) constitutes a group of primary vasculitides associated with antineutrophil cytoplasmic autoantibodies, which are either directed to proteinase-3 or myeloperoxidase. In contrast to other forms of vasculitis, immuohis

  2. ADP-glucose pyrophosphorylase is localized to both the cytoplasm and plastids in developing pericarp of tomato fruit

    Science.gov (United States)

    Chen, B. Y.; Wang, Y.; Janes, H. W.

    1998-01-01

    The intracellular location of ADP-glucose pyrophosphorylase (AGP) in developing pericarp of tomato (Lycopersicon esculentum Mill) has been investigated by immunolocalization. With the use of a highly specific anti-tomato fruit AGP antibody, the enzyme was localized in cytoplasm as well as plastids at both the light and electron microscope levels. The immunogold particles in plastids were localized in the stroma and at the surface of the starch granule, whereas those in the cytoplasm occurred in cluster-like patterns. Contrary to the fruit, the labeling in tomato leaf cells occurred exclusively in the chloroplasts. These data demonstrate that AGP is localized to both the cytoplasm and plastids in developing pericarp cells of tomato.

  3. Model for bidirectional movement of cytoplasmic dynein

    CERN Document Server

    Sumathy, S

    2014-01-01

    Cytoplasmic dynein exhibits a directional processive movement on microtubule filaments and is known to move in steps of varying length based on the number of ATP molecules bound to it and the load that it carries. It is experimentally observed that dynein takes occasional backward steps and the frequency of such backward steps increases as the load approaches the stall force. Using a stochastic process model, we investigate the bidirectional movement of single head of a dynein motor. The probability for backward step is implemented based on Crook's fluctuation theorem of non-equilibrium statistical mechanics. We find that the movement of dynein motor is characterized with negative velocity implying backward motion beyond stall force. We observe that the motor moves backward for super stall forces by hydrolyzing the ATP exactly the same way as it does while moving forward for sub stall forces.

  4. Inborn errors of cytoplasmic triglyceride metabolism.

    Science.gov (United States)

    Wu, Jiang Wei; Yang, Hao; Wang, Shu Pei; Soni, Krishnakant G; Brunel-Guitton, Catherine; Mitchell, Grant A

    2015-01-01

    Triglyceride (TG) synthesis, storage, and degradation together constitute cytoplasmic TG metabolism (CTGM). CTGM is mostly studied in adipocytes, where starting from glycerol-3-phosphate and fatty acyl (FA)-coenzyme A (CoA), TGs are synthesized then stored in cytoplasmic lipid droplets. TG hydrolysis proceeds sequentially, producing FAs and glycerol. Several reactions of CTGM can be catalyzed by more than one enzyme, creating great potential for complex tissue-specific physiology. In adipose tissue, CTGM provides FA as a systemic energy source during fasting and is related to obesity. Inborn errors and mouse models have demonstrated the importance of CTGM for non-adipose tissues, including skeletal muscle, myocardium and liver, because steatosis and dysfunction can occur. We discuss known inborn errors of CTGM, including deficiencies of: AGPAT2 (a form of generalized lipodystrophy), LPIN1 (childhood rhabdomyolysis), LPIN2 (an inflammatory condition, Majeed syndrome, described elsewhere in this issue), DGAT1 (protein loosing enteropathy), perilipin 1 (partial lipodystrophy), CGI-58 (gene ABHD5, neutral lipid storage disease (NLSD) with ichthyosis and "Jordan's anomaly" of vacuolated polymorphonuclear leukocytes), adipose triglyceride lipase (ATGL, gene PNPLA2, NLSD with myopathy, cardiomyopathy and Jordan's anomaly), hormone-sensitive lipase (HSL, gene LIPE, hypertriglyceridemia, and insulin resistance). Two inborn errors of glycerol metabolism are known: glycerol kinase (GK, causing pseudohypertriglyceridemia) and glycerol-3-phosphate dehydrogenase (GPD1, childhood hepatic steatosis). Mouse models often resemble human phenotypes but may diverge markedly. Inborn errors have been described for less than one-third of CTGM enzymes, and new phenotypes may yet be identified. PMID:25300978

  5. Inborn errors of cytoplasmic triglyceride metabolism.

    Science.gov (United States)

    Wu, Jiang Wei; Yang, Hao; Wang, Shu Pei; Soni, Krishnakant G; Brunel-Guitton, Catherine; Mitchell, Grant A

    2015-01-01

    Triglyceride (TG) synthesis, storage, and degradation together constitute cytoplasmic TG metabolism (CTGM). CTGM is mostly studied in adipocytes, where starting from glycerol-3-phosphate and fatty acyl (FA)-coenzyme A (CoA), TGs are synthesized then stored in cytoplasmic lipid droplets. TG hydrolysis proceeds sequentially, producing FAs and glycerol. Several reactions of CTGM can be catalyzed by more than one enzyme, creating great potential for complex tissue-specific physiology. In adipose tissue, CTGM provides FA as a systemic energy source during fasting and is related to obesity. Inborn errors and mouse models have demonstrated the importance of CTGM for non-adipose tissues, including skeletal muscle, myocardium and liver, because steatosis and dysfunction can occur. We discuss known inborn errors of CTGM, including deficiencies of: AGPAT2 (a form of generalized lipodystrophy), LPIN1 (childhood rhabdomyolysis), LPIN2 (an inflammatory condition, Majeed syndrome, described elsewhere in this issue), DGAT1 (protein loosing enteropathy), perilipin 1 (partial lipodystrophy), CGI-58 (gene ABHD5, neutral lipid storage disease (NLSD) with ichthyosis and "Jordan's anomaly" of vacuolated polymorphonuclear leukocytes), adipose triglyceride lipase (ATGL, gene PNPLA2, NLSD with myopathy, cardiomyopathy and Jordan's anomaly), hormone-sensitive lipase (HSL, gene LIPE, hypertriglyceridemia, and insulin resistance). Two inborn errors of glycerol metabolism are known: glycerol kinase (GK, causing pseudohypertriglyceridemia) and glycerol-3-phosphate dehydrogenase (GPD1, childhood hepatic steatosis). Mouse models often resemble human phenotypes but may diverge markedly. Inborn errors have been described for less than one-third of CTGM enzymes, and new phenotypes may yet be identified.

  6. The epididymis, cytoplasmic droplets and male fertility

    Institute of Scientific and Technical Information of China (English)

    Trevor G Cooper

    2011-01-01

    The potential of spermatozoa to become motile during post-testicular maturation,and the relationship between the cytoplasmic droplet and fertilizing capacity are reviewed.Post-testicular maturation of spermatozoa involves the autonomous induction of motility,which can occur in vivo in testes with occluded excurrent ducts and in vitro in testicular explants,and artefactual changes in morphology that appear to occur in the testis in vitro.Both modifications may reflect time-dependent oxidation of disulphide bonds of head and tail proteins.Regulatory volume decrease(RVD),which counters sperm swelling at ejaculation,is discussed in relation to loss of cytoplasmic droplets and consequences for fertility.It is postulated that:(i)fertile males possess spermatozoa with sufficient osmolytes to drive RVD at ejaculation,permitting the droplet to round up and pinch off without membrane rupture; and(ⅱ)infertile males possess spermatozoa with insufficient osmolytes so that RVD is inadequate,the droplet swells and the resulting flagellar angulation prevents droplet loss.Droplet retention at ejaculation is a harbinger of infertility caused by failure of the spermatozoon to negotiate the uterotubal junction or mucous and reach the egg.In this hypothesis,the epididymis regulates fertility indirectly by the extent of osmolyte provision to spermatozoa,which influences RVD and therefore droplet loss.Man is an exception,because ejaculated human spermatozoa retain their droplets.This may reflect their short midpiece,approximating head length,permitting a swollen droplet to extend along the entire midpiece; this not only obviates droplet migration and flagellar angulation but also hampers droplet loss.

  7. The molecular mechanism and physiological role of cytoplasmic streaming.

    Science.gov (United States)

    Tominaga, Motoki; Ito, Kohji

    2015-10-01

    Cytoplasmic streaming occurs widely in plants ranging from algae to angiosperms. However, the molecular mechanism and physiological role of cytoplasmic streaming have long remained unelucidated. Recent molecular genetic approaches have identified specific myosin members (XI-2 and XI-K as major and XI-1, XI-B, and XI-I as minor motive forces) for the generation of cytoplasmic streaming among 13 myosin XIs in Arabidopsis thaliana. Simultaneous knockout of these myosin XI members led to a reduced velocity of cytoplasmic streaming and marked defects of plant development. Furthermore, the artificial modifications of myosin XI-2 velocity changed plant and cell sizes along with the velocity of cytoplasmic streaming. Therefore, we assume that cytoplasmic streaming is one of the key regulators in determining plant size.

  8. Purification and properties of the cytoplasmic glucose-6-phosphate dehydrogenase from pea leaves.

    Science.gov (United States)

    Fickenscher, K; Scheibe, R

    1986-06-01

    A method involving affinity chromatography on the yellow dye Remazol Brilliant Gelb GL to highly purify the cytoplasmic isoenzyme of glucose-6-phosphate dehydrogenase from pea shoots is described. Purification is at least 6000-fold. The specific activity of the purified enzyme is 185 mumol NADP reduced/min per mg protein. The preparation was free from any contamination of chloroplastic isoenzyme. The purified enzyme retains its activity in the presence of reducing agents which, in contrast, inactivate the chloroplast enzyme. The state of activity of the cytoplasmic and the chloroplastic isoenzyme in illuminated or darkened pea leaves was investigated using specific antibodies. While upon illumination the chloroplastic isoenzyme was inactivated by 80 to 90%, we could not find any change in activity of the cytoplasmic glucose-6-phosphate dehydrogenase. ATP, ADP, NAD, NADH, and various sugar phosphates do not inhibit the enzyme activity. Only NADPH is a strong competitive inhibitor with respect to NADP, suggesting that the enzyme is regulated by feedback inhibition by one of its products. Mg2+ ions have no influence on the activity of the enzyme. The molecular weight has found to be 240,000 for the native enzyme and 60,000 for the subunit. Throughout the purification procedure the enzyme was very unstable unless NADP was present in the buffer. PMID:3717951

  9. A universal strategy for stable intracellular antibodies.

    Science.gov (United States)

    Shaki-Loewenstein, Shelly; Zfania, Rahely; Hyland, Stephen; Wels, Winfried S; Benhar, Itai

    2005-08-01

    The expression of intracellular antibodies (intrabodies) in mammalian cells has provided a powerful tool to manipulate microbial and cellular signalling pathways in a highly precise manner. However, several technical hurdles have thus far restricted their more widespread use. In particular, single-chain antibodies (scFvs) have been reported to fold poorly in the reducing environment of the cytoplasm and as such there has been a reluctance to use scFv-phage libraries as a source of intrabodies unless a preselection step was applied to identify these rare scFvs that could fold properly in the absence of disulfide bonds. Recently, we reported that scFvs can be efficiently expressed within the cytoplasm of bacteria when fused at the C-terminus of the Escherichia coli maltose-binding protein (MBP). Here, we demonstrate that such MBP-scFvs are similarly stabilized when expressed in the mammalian cell cytoplasm as well as other compartments. This was demonstrated by comparing MBP-scFv fusions to the corresponding unfused scFvs that activate a defective beta-galactosidase enzyme, others that neutralize the wild-type beta-galactosidase enzyme, and an antibody that blocks the epidermal growth factor receptor. In all cases, the MBP-scFvs significantly outperformed their unfused counterparts. Our results suggest that fusion of scFvs to MBP, and possibly to other "chaperones in the context of a fusion protein", may provide a universal approach for efficient expression of intrabodies in the mammalian cell cytoplasm. This strategy should allow investigators to bypass much of the in vitro scFv characterization that is often not predictive of in vivo intrabody function and provide a more efficient use of large native and synthetic scFv-phage libraries already in existence to identify intrabodies that will be active in vivo.

  10. Impairment and Differential Expression of PR3 and MPO on Peripheral Myelomonocytic Cells with Endothelial Properties in Granulomatosis with Polyangiitis

    OpenAIRE

    Susann Patschan; Daniel Patschan; Elvira Henze; Sabine Blaschke; Wessels, Johannes T.; Gerhard Anton Müller

    2012-01-01

    Background. Granulomatosis with polyangiitis (GPA) and microscopic polyangiitis (MPA) are autoimmune-mediated diseases characterized by vasculitic inflammation of respiratory tract and kidneys. Clinical observations indicated a strong association between disease activity and serum levels of certain types of autoantibodies (antineutrophil cytoplasm antibodies with cytoplasmic [cANCA in GPA] or perinuclear [pAN CA in MPA] immunofluorescence). Pathologically, both diseases are characterized by s...

  11. [Pulmonar pseudotumor in granulomatosis with polyangiitis (GPA). Pulmonary cancer and/or GPA? Diagnostic implications of pulmonary nodules].

    Science.gov (United States)

    Horta-Baas, Gabriel; Meza-Zempoaltecatl, Esteban; Pérez-Cristóbal, Mario; Barile-Fabris, Leonor Adriana

    2016-01-01

    Granulomatosis with polyangiitis (GPA), formerly known as Wegener's granulomatosis, is a systemic necrotizing vasculitis, which affects small and medium sized blood vessels and is often associated with cytoplasmic anti-neutrophil cytoplasmic antibodies (ANCA). Inflammatory pseudotumor is a rare condition characterized by the appearance of a mass lesion that mimics a malignant tumor both clinically and on imaging studies, but that is thought to have an inflammatory/reactive pathogenesis. We report a patient with a GPA which was originally diagnosed as malignancy.

  12. Granulomatosis with polyangiitis and associated pulmonary emphysema: Breathtaking vasculitis

    OpenAIRE

    Gadre, Shruti K.; Stoller, James K; Mehta, Atul C.

    2015-01-01

    Pulmonary emphysema occasionally occurs in the absence of smoking or noxious exposures. Other than through a known association with alpha-1 antitryspin deficiency, to our knowledge, no reports implicate granulomatosis with polyangiitis (GPA) in causing airflow obstruction with small airway involvement and severe air trapping. To extend available experience, we report a 51-year-old male with biopsy-proven cytoplasmic-antineutrophilic cytoplasmic antibody proteinase 3 (ANCA PR3)-positive GPA wh...

  13. Mitochondrial DNA Phylogeny of Cultivated and Wild Beets: Relationships Among Cytoplasmic Male-Sterility-Inducing and Nonsterilizing Cytoplasms

    OpenAIRE

    Nishizawa, Satsuki; Mikami, Tetsuo; Kubo, Tomohiko

    2007-01-01

    Cytoplasmic male sterility (CMS), the maternally inherited failure to produce functional pollen, has been used in the breeding of sugar beet (Beta vulgaris ssp. vulgaris). At least three different sources of CMS can be distinguished from one another as well as from normal fertile cytoplasm by polymorphisms in their mitochondrial genomes. Here we analyzed 50 accessions of cultivated and wild beets to investigate the phylogenetic relationships among male-sterility-inducing and normal cytoplasms...

  14. Cytoplasmic Dynein Promotes HIV-1 Uncoating

    Directory of Open Access Journals (Sweden)

    Paulina Pawlica

    2014-11-01

    Full Text Available Retroviral capsid (CA cores undergo uncoating during their retrograde transport (toward the nucleus, and/or after reaching the nuclear membrane. However, whether HIV-1 CA core uncoating is dependent upon its transport is not understood. There is some evidence that HIV-1 cores retrograde transport involves cytoplasmic dynein complexes translocating on microtubules. Here we investigate the role of dynein-dependent transport in HIV-1 uncoating. To interfere with dynein function, we depleted dynein heavy chain (DHC using RNA interference, and we over-expressed p50/dynamitin. In immunofluorescence microscopy experiments, DHC depletion caused an accumulation of CA foci in HIV-1 infected cells. Using a biochemical assay to monitor HIV-1 CA core disassembly in infected cells, we observed an increase in amounts of intact (pelletable CA cores upon DHC depletion or p50 over-expression. Results from these two complementary assays suggest that inhibiting dynein-mediated transport interferes with HIV-1 uncoating in infected cells, indicating the existence of a functional link between HIV-1 transport and uncoating.

  15. A physical perspective on cytoplasmic streaming.

    Science.gov (United States)

    Goldstein, Raymond E; van de Meent, Jan-Willem

    2015-08-01

    Organisms show a remarkable range of sizes, yet the dimensions of a single cell rarely exceed 100 µm. While the physical and biological origins of this constraint remain poorly understood, exceptions to this rule give valuable insights. A well-known counterexample is the aquatic plant Chara, whose cells can exceed 10 cm in length and 1 mm in diameter. Two spiralling bands of molecular motors at the cell periphery drive the cellular fluid up and down at speeds up to 100 µm s(-1), motion that has been hypothesized to mitigate the slowness of metabolite transport on these scales and to aid in homeostasis. This is the most organized instance of a broad class of continuous motions known as 'cytoplasmic streaming', found in a wide range of eukaryotic organisms-algae, plants, amoebae, nematodes and flies-often in unusually large cells. In this overview of the physics of this phenomenon, we examine the interplay between streaming, transport and cell size and discuss the possible role of self-organization phenomena in establishing the observed patterns of streaming.

  16. A physical perspective on cytoplasmic streaming (invited)

    CERN Document Server

    Goldstein, Raymond E

    2015-01-01

    Organisms show a remarkable range of sizes, yet the dimensions of a single cell rarely exceed $100$ $\\mu$m. While the physical and biological origins of this constraint remain poorly understood, exceptions to this rule give valuable insights. A well-known counterexample is the aquatic plant $Chara$, whose cells can exceed $10$ cm in length and $1$ mm in diameter. Two spiraling bands of molecular motors at the cell periphery drive the cellular fluid up and down at speeds up to $100$ $\\mu$m/s, motion that has been hypothesized to mitigate the slowness of metabolite transport on these scales and to aid in homeostasis. This is the most organized instance of a broad class of continuous motions known as "cytoplasmic streaming", found in a wide range of eukaryotic organisms - algae, plants, amoebae, nematodes, and flies - often in unusually large cells. In this overview of the physics of this phenomenon, we examine the interplay between streaming, transport and cell size, and discuss the possible role of self-organi...

  17. Single molecule analysis of cytoplasmic dynein motility

    Science.gov (United States)

    Yildiz, Ahmet

    2014-03-01

    Cytoplasmic dynein is a homodimeric AAA + motor that transports a multitude of cargos towards the microtubule (MT) minus end. The mechanism of dynein motility remains unclear, due to its large size (2.6 MDa) and the complexity of its structure. By tracking the stepping motion of both heads at nanometer resolution, we observed that dynein heads move independently along the MT, in contrast to hand over hand movement of kinesins and myosin. Stepping behavior of the heads varies as a function of interhead separation and establishing the basis of high variability in dynein step size. By engineering the mechanical and catalytic properties of the dynein motor domain, we show that a rigid linkage between monomers and dimerization between N-terminal tail domains are not essential for processive movement. Instead, dynein processivity minimally requires the linker domain of one active monomer to be attached to an inert MT tether retaining only the MT-binding domain. The release of a dynein monomer from the MT can be mediated either by nucleotide binding or external load. Nucleotide dependent release is inhibited by the tension on the linker domain at high interhead separations. Tension dependent release is highly asymmetric, with faster release towards the minus-end. Reversing the asymmetry of the MT binding interface results in plus end directed motility, even though the force was generated by the dynein motor activity. On the basis of these measurements, we propose a model that describes the basis of dynein processivity, directionality and force generation.

  18. Developing improved durum wheat germplasm by altering the cytoplasmic genome

    Science.gov (United States)

    In eukaryotic organisms, nuclear and cytoplasmic genomes interact to drive cellular functions. These genomes have co-evolved to form specific nuclear-cytoplasmic interactions that are essential to the origin, success, and evolution of diploid and polyploid species. Hundreds of genetic diseases in h...

  19. Monoclonal antibodies

    International Nuclear Information System (INIS)

    Monoclonal antibodies (MAbs) are antibodies having single specificity for a given antigen site (epitope). The development of hybridoma technology and the relative ease by which MAbs can be prepared has revolutionized many aspects of serological applications in diagnosis and differentiation of disease producing agents. The property of monospecificity offers advantages in diagnostic applications over polyclonal sera in that tests can be defined exactly with regard to the antigen detected and the affinity of reaction between the given antigenic site and the monoclonal reagent. In addition, MAbs offer better possibilities for test standardization, because the same reagent can be used in different laboratories. Such an MAb can be supplied by a central laboratory or 'grown' from hybridoma cells, ensuring that the resultant product is identical from laboratory to laboratory and that the part of the test involving the MAb reaction is the same. The methodologies for inoculation regimes, mice, cloning methods, selection of fusion partners, etc., have been validated extensively in developed country laboratories. The decision to establish a MAb production facility must be examined on a strict cost-benefit basis, since it is still expensive to produce a product. There are many MAbs available that should be sought to allow exploitation in developing tests. If a production facility is envisaged, it should produce reagents for national needs, i.e. there should be a clear problem oriented approach whereby exact needs are defined. In the field of veterinary applications, MAbs are the central reagent in many immunoassays based on the enzyme linked immunosorbent assay (ELISA). The development of specific tests for diagnosing diseases is dominated by MAbs and has been fuelled by a strong research base, mainly in developed countries allied to developing countries through the study of related diseases. Thus, there are very many assays dependent on MAbs, some of which form the basis of

  20. Cytoplasmic streaming velocity as a plant size determinant.

    Science.gov (United States)

    Tominaga, Motoki; Kimura, Atsushi; Yokota, Etsuo; Haraguchi, Takeshi; Shimmen, Teruo; Yamamoto, Keiichi; Nakano, Akihiko; Ito, Kohji

    2013-11-11

    Cytoplasmic streaming is active transport widely occurring in plant cells ranging from algae to angiosperms. Although it has been revealed that cytoplasmic streaming is generated by organelle-associated myosin XI moving along actin bundles, the fundamental function in plants remains unclear. We generated high- and low-speed chimeric myosin XI by replacing the motor domains of Arabidopsis thaliana myosin XI-2 with those of Chara corallina myosin XI and Homo sapiens myosin Vb, respectively. Surprisingly, the plant sizes of the transgenic Arabidopsis expressing high- and low-speed chimeric myosin XI-2 were larger and smaller, respectively, than that of the wild-type plant. This size change correlated with acceleration and deceleration, respectively, of cytoplasmic streaming. Our results strongly suggest that cytoplasmic streaming is a key determinant of plant size. Furthermore, because cytoplasmic streaming is a common system for intracellular transport in plants, our system could have applications in artificial size control in plants.

  1. Long Non-coding RNAs in the Cytoplasm

    Institute of Scientific and Technical Information of China (English)

    Farooq Rashid; Abdullah Shah; Ge Shan

    2016-01-01

    An enormous amount of long non-coding RNAs (lncRNAs) transcribed from eukaryotic genome are important regulators in different aspects of cellular events. Cytoplasm is the residence and the site of action for many lncRNAs. The cytoplasmic lncRNAs play indispensable roles with multiple molecular mechanisms in animal and human cells. In this review, we mainly talk about functions and the underlying mechanisms of lncRNAs in the cytoplasm. We highlight relatively well-studied examples of cytoplasmic lncRNAs for their roles in modulating mRNA stability, regulating mRNA translation, serving as competing endogenous RNAs, functioning as precursors of microRNAs, and mediating protein modifications. We also elaborate the perspectives of cytoplasmic lncRNA studies.

  2. Serine phosphorylation of syndecan-2 proteoglycan cytoplasmic domain

    DEFF Research Database (Denmark)

    Oh, E S; Couchman, J R; Woods, A

    1997-01-01

    Protein kinase C (PKC) is involved in cell-matrix and cell-cell adhesion, and the cytoplasmic domain of syndecan-2 contains two serines (residues 197 and 198) which lie in a consensus sequence for phosphorylation by PKC. Other serine and threonine residues are present but not in a consensus...... sequence. We investigated phosphorylation of syndecan-2 cytoplasmic domain by PKC, using purified GST-syndecan-2 fusion proteins and synthetic peptides corresponding to regions of the cytoplasmic domain. A synthetic peptide encompassing the entire cytoplasmic domain of syndecan-2 was phosphorylated by PKC...... cytoplasmic domain peptides were monomeric, with 2 mol/mol serine phosphorylation. At higher concentrations, however, the peptides formed dimers, with only 0.5 mol/mol phosphorylation. Concentration-dependent dimerization was not altered by phosphorylation. Phosphorylation is, therefore, dependent on the...

  3. Monoclonal antibodies.

    Science.gov (United States)

    2009-01-01

    The ability to produce and exploit monoclonal antibodies (mAbs) has revolutionized many areas of biological sciences. The unique property of an mAb is that it is a single species of immunoglobulin (IG) molecule. This means that the specificity of the interaction of the paratopes on the IG, with the epitopes on an antigenic target, is the same on every molecule. This property can be used to great benefit in immunoassays to provide tests of defined specificity and sensitivity, which improve the possibilities of standardization. The performance of assays can often be determined relating the actual weight of antibody (hence the number of molecules) to the activity. Often the production of an mAb against a specific epitope is the only way that biological entities can be differentiated. This chapter outlines the areas involving the development of assays based on mAbs. The problems involved address include the physical aspects of mAbs and how they may affect assay design and also the implications of results based on monospecific reagents. Often these are not fully understood, leading to assays that are less than satisfactory, which does not justify the relatively high cost of preparing and screening of mAbs. There are many textbooks and reviews dealing with the preparation of mAbs, the principles involved, and various purification and manipulative methods for the preparation of fragments and conjugation. There has been little general information attempting to summarize the best approaches to assay design using mAbs. Much time can be wasted through bad planning, and this is particularly relevant to mAbs. A proper understanding of some basic principles is essential. It is beyond the scope of this chapter to discuss all aspects, but major areas are highlighted. PMID:19219589

  4. CTP synthase forms cytoophidia in the cytoplasm and nucleus

    Energy Technology Data Exchange (ETDEWEB)

    Gou, Ke-Mian [MRC Functional Genomics Unit, Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford OX1 3PT (United Kingdom); State Key Laboratory for Agrobiotechnology, College of Biological Sciences, China Agricultural University, Beijing 100193 (China); Chang, Chia-Chun [Institute of Biotechnology, National Taiwan University, Taipei, Taiwan, ROC (China); Shen, Qing-Ji [MRC Functional Genomics Unit, Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford OX1 3PT (United Kingdom); Sung, Li-Ying, E-mail: liyingsung@ntu.edu.tw [Institute of Biotechnology, National Taiwan University, Taipei, Taiwan, ROC (China); Agricultural Biotechnology Research Center, Academia Sinica, Taipei 115, Taiwan, ROC (China); Liu, Ji-Long, E-mail: jilong.liu@dpag.ox.ac.uk [MRC Functional Genomics Unit, Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford OX1 3PT (United Kingdom)

    2014-04-15

    CTP synthase is an essential metabolic enzyme responsible for the de novo synthesis of CTP. Multiple studies have recently showed that CTP synthase protein molecules form filamentous structures termed cytoophidia or CTP synthase filaments in the cytoplasm of eukaryotic cells, as well as in bacteria. Here we report that CTP synthase can form cytoophidia not only in the cytoplasm, but also in the nucleus of eukaryotic cells. Both glutamine deprivation and glutamine analog treatment promote formation of cytoplasmic cytoophidia (C-cytoophidia) and nuclear cytoophidia (N-cytoophidia). N-cytoophidia are generally shorter and thinner than their cytoplasmic counterparts. In mammalian cells, both CTP synthase 1 and CTP synthase 2 can form cytoophidia. Using live imaging, we have observed that both C-cytoophidia and N-cytoophidia undergo multiple rounds of fusion upon glutamine analog treatment. Our study reveals the coexistence of cytoophidia in the cytoplasm and nucleus, therefore providing a good opportunity to investigate the intracellular compartmentation of CTP synthase. - Highlights: • CTP synthase forms cytoophidia not only in the cytoplasm but also in the nucleus. • Glutamine deprivation and Glutamine analogs promotes cytoophidium formation. • N-cytoophidia exhibit distinct morphology when compared to C-cytoophidia. • Both CTP synthase 1 and CTP synthase 2 form cytoophidia in mammalian cells. • Fusions of cytoophidia occur in the cytoplasm and nucleus.

  5. Differences in catalytic properties between cerebral cytoplasmic and mitochondrial hexokinases.

    Science.gov (United States)

    Thompson, M F; Bachelard, H S

    1977-03-01

    1. Clear kinetic differences between cytoplasmic and mitochondrial forms of type-I cerebral hexokinase were demonstrated from experiments performed under identical conditions on three (cytoplasmic, bound mitochondrial and solubilized mitochondrial) preparations of the enzyme. 2. Whereas the Michaelis constant for glucose (KmGlc) was consistent, that for MgATP2- (KmATP) was lower in the cytoplasmic than in the two mitochondrial preparations. The substrate dissociation constants (KsGlc and KsATP) were both higher in the cytoplasmic than in the mitochondrial preparations. A further difference in the substrate kinetic patterns was that KmATP=KmATP for the cytoplasmic enzyme, in contrast with the mitochondrial enzyme, where KmATP was clearly not equal to KsATP [Bachelard et al. (1971) Biochem. J. 123, 707-715]. 3. Dead-end inhibition produced by N-acetyl-glucosamine and by AMP also exhibited different quantitative kinetic patterns for the two enzyme sources. Both inhibitions gave Ki values similar or equal to those of Ki' for the cytoplasmic activity, whereas Ki was clearly not equal to Ki' for the mitochondrial activity. 4. All of these studies demonstrated the similarity of the two mitochondrial activities (particulate and solubilized), which were both clearly different from the cytoplasmic activity. 5. The analysis gives a practical example of our previous theoretical treatment on the derivation of true inhibition constants. 6. The results are discussed in terms of the function of cerebral hexokinases.

  6. A young lady presented with limited pulmonary Wegener′s granulomatosis

    Directory of Open Access Journals (Sweden)

    Dey Atin

    2008-01-01

    Full Text Available A 19 year old female college student presented with fever, dry cough, chest pain, blood tinged sputum with subsequent development of polyarthralgia with radiological evidence of bilateral multiple unevenly distributed pulmonary nodular opacities with cavitation. There was no other systemic involvement and the patient was cytoplasmic antineutrophil antibody (c-ANCA positive with more than four times the normal upper limit of anti PR3 antibody. Excellent response to oral steroid with antimicrobial agent "trimethoprim - sulphamethoxazole" was noted.

  7. The rise and fall of horror autotoxicus and forbidden clones.

    Science.gov (United States)

    Jennette, J Charles; Falk, Ronald J

    2010-09-01

    Cui and associates show that healthy individuals have natural autoantibodies (NAAs) specific for myeloperoxidase, proteinase 3, and glomerular basement membrane (GBM) with the same specificity as anti-neutrophil cytoplasmic antibodies and anti-GBM antibodies that are pathogenic. Although Ehrlich proposed horror autotoxicus and Burnet envisioned elimination of forbidden clones, NAAs are present in all healthy individuals and play beneficial homeostatic roles. Pathogenic autoimmunity is dysregulation of natural homeostatic autoimmunity rather than onset of a previously absent self-recognition.

  8. Cost-effective detection of non-antidouble-stranded DNA antinuclear antibody specificities in daily clinical practice

    NARCIS (Netherlands)

    Vos, PAJM; Bast, EJEG; Derksen, RHWM

    2006-01-01

    Objectives. To compare the utility of indirect immunofluorescence for the detection of antinuclear antibodies (ANA-IIF) and a fully automated test (ELiA Symphony (TM)) that detects antibodies against a mixture of nuclear and cytoplasmic antigens (ENA), to select sera that should be tested for non-an

  9. Adaptive responses to antibody based therapy.

    Science.gov (United States)

    Rodems, Tamara S; Iida, Mari; Brand, Toni M; Pearson, Hannah E; Orbuch, Rachel A; Flanigan, Bailey G; Wheeler, Deric L

    2016-02-01

    Receptor tyrosine kinases (RTKs) represent a large class of protein kinases that span the cellular membrane. There are 58 human RTKs identified which are grouped into 20 distinct families based upon their ligand binding, sequence homology and structure. They are controlled by ligand binding which activates intrinsic tyrosine-kinase activity. This activity leads to the phosphorylation of distinct tyrosines on the cytoplasmic tail, leading to the activation of cell signaling cascades. These signaling cascades ultimately regulate cellular proliferation, apoptosis, migration, survival and homeostasis of the cell. The vast majority of RTKs have been directly tied to the etiology and progression of cancer. Thus, using antibodies to target RTKs as a cancer therapeutic strategy has been intensely pursued. Although antibodies against the epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER2) have shown promise in the clinical arena, the development of both intrinsic and acquired resistance to antibody-based therapies is now well appreciated. In this review we provide an overview of the RTK family, the biology of EGFR and HER2, as well as an in-depth review of the adaptive responses undertaken by cells in response to antibody based therapies directed against these receptors. A greater understanding of these mechanisms and their relevance in human models will lead to molecular insights in overcoming and circumventing resistance to antibody based therapy. PMID:26808665

  10. Regulation of cytoplasmic streaming in Vallisneria mesophyll cells.

    Science.gov (United States)

    Takagi, S; Nagai, R

    1983-07-01

    Induction and cessation of the rotational cytoplasmic streaming in Vallisneria mesophyll cells could be controlled by external stimuli. In cells that had been kept in darkness the cytoplasm remained quiescent. However, when the cells were treated in the dark with EGTA solution (10 mM or 20 mM buffered with 10 mM-Tris-maleate at pH 7.0), rotational cytoplasmic streaming was induced. When the cells were transferred again to artificial pond water in the dark, the induced streaming was inhibited; that is, only 50% of the observed cells exhibited active streaming after 2 h. When the cells were irradiated continuously with far-red light (lambda max = 750 nm, 0.4 W/m2) in the same external medium, the induced streaming was inhibited almost completely within 2 h. The relative quantum effectiveness of monochromatic light (450--800 nm) in producing cessation of streaming was also investigated. Irradiation with light of 450, 550 and 600 nm was almost as effective as darkness. Light of 500 and 650 nm was less effective than dark exposure. Only irradiation at 750 nm stopped streaming in almost all cells. But when calcium was excluded from the external medium, the effect of far-red light decreased to almost the dark control level. Light of 800 nm also inhibited the streaming but the effect was much less than that of far-red light. Microfilaments in bundles with the long axis parallel to the streaming direction were localized in the vicinity of the cell membrane. Their configuration, localization and distribution were the same in the present experimental system irrespective of whether the cytoplasm was streaming or quiescent. Intracellular calcium was examined by electron microscopic cytochemistry and X-ray microanalysis. In cells with streaming induced by EGTA, only a small amount of calcium-containing precipitates formed in the cytoplasm in the presence of antimony. A few precipitates were found in the chloroplasts, the middle lamella of the cell wall and at the border between

  11. Activated caspase-9 immunoreactivity in glial and neuronal cytoplasmic inclusions in multiple system atrophy.

    Science.gov (United States)

    Kawamoto, Yasuhiro; Ayaki, Takashi; Urushitani, Makoto; Ito, Hidefumi; Takahashi, Ryosuke

    2016-08-15

    The mitochondria play an important role in apoptotic cell death, and the released cytochrome c from the mitochondria promotes the formation of the apoptosome, which contains cytochrome c, Apaf-1 and caspase-9, resulting in the activation of caspase-9 and the promotion of the apoptotic cascade. To investigate the role of mitochondria-dependent apoptotic cell death in patients with multiple system atrophy (MSA), we performed immunohistochemical studies on apoptosome-related proteins in formalin-fixed, paraffin-embedded sections from 8 normal subjects and 10 patients with MSA. We then performed double-labeling immunohistochemistry for activated caspase-9 and α-synuclein in some sections from 10 patients with MSA. In the brains with MSA, glial cytoplasmic inclusions (GCIs) and neuronal cytoplasmic inclusions (NCIs) were intensely immunoreactive for cytochrome c, Apaf-1 and caspase-9. Activated caspase-9 immunoreactivities were also confirmed to be densely localized to both GCIs and NCIs using two types of anti-cleaved caspase-9 antibodies. The semiquantitative analyses using the upper pontine sections double-immunostained with cleaved caspase-9 and α-synuclein demonstrated that approximately 80% of GCIs and NCIs were immunopositive for cleaved caspase-9. Our results suggest that the formation of the apoptosome accompanied by the activation of caspase-9 may occur in brains affected by MSA, and that a mitochondria-dependent apoptotic pathway may be partially associated with the pathogenesis of MSA. PMID:27345387

  12. Cytoplasmic Localization of Human cdc25C during Interphase Requires an Intact 14-3-3 Binding Site

    OpenAIRE

    Dalal, Sorab N.; Schweitzer, Colleen M.; Gan, Jianmin; DeCaprio, James A.

    1999-01-01

    cdc25C induces mitosis by activating the cdc2-cyclin B complex. The intracellular localization of cyclin B1 is regulated in a cell cycle-specific manner, and its entry into the nucleus may be required for the initiation of mitosis. To determine the cellular localization of cdc25C, monoclonal antibodies specific for cdc25C were developed and used to demonstrate that in human cells, cdc25C is retained in the cytoplasm during interphase. A deletion analysis identified a 58-amino-acid region (ami...

  13. A new tool for plant cell biology: in vivo antibody uptake in plant protoplasts.

    Science.gov (United States)

    Brière, C; Barthou, H; Petitprez, M

    2004-07-01

    We report on the in vivo uptake of antibodies into plant protoplasts. When protoplasts of sunflower, Arabidopsis or tobacco were incubated in vivo with an antibody, this antibody was detected by immunofluorescence in the cytoplasm and/or the nucleus, depending on the location of the target protein. Furthermore, when protoplasts were cultured in the presence of antibodies, specific effects were observed. Incubation with antibodies raised against p34cdc2 led to a strong inhibition of the division rate, and a decrease in the average DNA content of protoplasts. With antibodies against HaWLIM1, a LIM domain protein of the CRP type, a negative effect on actin organisation was observed. We conclude that antibodies can penetrate plant protoplasts in vivo, and thus may be used as powerful tools for the study of protein function.

  14. Monoclonal antibodies against plant proteins recognise animal intermediate filaments.

    Science.gov (United States)

    Parke, J M; Miller, C C; Cowell, I; Dodson, A; Dowding, A; Downes, M; Duckett, J G; Anderton, B J

    1987-01-01

    Four monoclonal antibodies were raised against polypeptides present in a high-salt detergent-insoluble fraction from cells of Chlamydomonas reinhardtii. Indirect immunofluorescence microscopy of fibroblasts and epithelial cells grown in culture using these plant antibodies revealed staining arrays identical to those obtained with well characterised antibodies to animal intermediate filaments. Immunofluorescence microscopy of Chlamydomonas with these monoclonal antibodies and a monoclonal antibody that recognises all animal intermediate filaments (anti-IFA) gave a diffuse, patchy cytoplasmic staining pattern. Both the plant antibodies and anti-IFA stained interphase onion root tip cells in a diffuse perinuclear pattern. In metaphase through to telophase, the labelling patterns colocalised with those of microtubules. Labelling of the phragmoplast was also detected but not staining of the preprophase band. On Western blots of various animal cell lines and tissues, all the antibodies labelled known intermediate filament proteins. On Western blots of whole Chlamydomonas proteins, all the antibodies labelled a broad band in the 57,000 Mr range, and three antibodies labelled bands around 66,000 and 140,000 Mr but with variable intensities. On Western blots of whole onion root tip proteins, all the antibodies labelled 50,000 Mr (two to three bands) polypeptides and a diffuse band around 60,000 Mr and three of the antibodies also labelled several polypeptides in the 90,000-200,000 Mr range. The consistent labelling of these different bands by several different monoclonal antibodies recognising animal intermediate filaments makes these polypeptides putative plant intermediate filament proteins. PMID:2446785

  15. Development of Biodegradable Nanocarriers Loaded with a Monoclonal Antibody

    Directory of Open Access Journals (Sweden)

    Andrew Gdowski

    2015-02-01

    Full Text Available Treatments utilizing monoclonal antibody therapeutics against intracellular protein-protein interactions in cancer cells have been hampered by several factors, including poor intracellular uptake and rapid lysosomal degradation. Our current work examines the feasibility of encapsulating monoclonal antibodies within poly(lactic-co-glycolic acid (PLGA nanoparticles using a water/oil/water double emulsion solvent evaporation technique. This method can be used to prepare protective polymeric nanoparticles for transporting functional antibodies to the cytoplasmic compartment of cancer cells. Nanoparticles were formulated and then characterized using a number of physical and biological parameters. The average nanoparticle size ranged from 221 to 252 nm with a low polydispersity index. Encapsulation efficiency of 16%–22% and antibody loading of 0.3%–1.12% were observed. The antibody molecules were released from the nanoparticles in a sustained manner and upon release maintained functionality. Our studies achieved successful formulation of antibody loaded polymeric nanoparticles, thus indicating that a PLGA-based antibody nanoformulation is a promising intracellular delivery vehicle for a large number of new intracellular antibody targets in cancer cells.

  16. Neutralizing Antibodies and Persistence of Immunity following Anthrax Vaccination

    OpenAIRE

    Hanson, James F.; Taft, Sarah C.; Weiss, Alison A.

    2006-01-01

    Anthrax toxin consists of protective antigen (PA) and two toxic components, lethal factor (LF) and edema factor (EF). PA binds to mammalian cellular receptors and delivers the toxic components to the cytoplasm. PA is the primary antigenic component of the current anthrax vaccine. Immunity is due to the generation of antibodies that prevent the PA-mediated internalization of LF and EF. In this study, we characterized sera obtained from vaccinated military personnel. Anthrax vaccine is administ...

  17. Mitochondrial DNA phylogeny of cultivated and wild beets: relationships among cytoplasmic male-sterility-inducing and nonsterilizing cytoplasms.

    Science.gov (United States)

    Nishizawa, Satsuki; Mikami, Tetsuo; Kubo, Tomohiko

    2007-11-01

    Cytoplasmic male sterility (CMS), the maternally inherited failure to produce functional pollen, has been used in the breeding of sugar beet (Beta vulgaris ssp. vulgaris). At least three different sources of CMS can be distinguished from one another as well as from normal fertile cytoplasm by polymorphisms in their mitochondrial genomes. Here we analyzed 50 accessions of cultivated and wild beets to investigate the phylogenetic relationships among male-sterility-inducing and normal cytoplasms. The haplotypes were characterized by the nucleotide sequence of the mitochondrial cox2-cox1 spacer region and mitochondrial minisatellite loci. The results indicated that (1) a normal cytoplasm line, cv. TK81-O, was situated at the major core node of the haplotype network, and (2) the three sterilizing cytoplasms in question derived independently from the core haplotype. The evolutionary pathway was investigated by physical mapping study of the mitochondrial genome of a wild beet (B. vulgaris ssp. orientalis) accession BGRC56777 which shared the same mitochondrial haplotype with TK81-O, but was not identical to TK81-O for the RFLP profiles of mitochondrial DNA. Interestingly, three sets of inverted repeated sequences appeared to have been involved in a series of recombination events during the course of evolution between the BGRC56777 and the TK81-O mitochondrial genomes.

  18. Progesterone influences cytoplasmic maturation in porcine oocytes developing in vitro.

    Science.gov (United States)

    Yuan, Bao; Liang, Shuang; Jin, Yong-Xun; Kwon, Jeong-Woo; Zhang, Jia-Bao; Kim, Nam-Hyung

    2016-01-01

    Progesterone (P4), an ovarian steroid hormone, is an important regulator of female reproduction. In this study, we explored the influence of progesterone on porcine oocyte nuclear maturation and cytoplasmic maturation and development in vitro. We found that the presence of P4 during oocyte maturation did not inhibit polar body extrusions but significantly increased glutathione and decreased reactive oxygen species (ROS) levels relative to that in control groups. The incidence of parthenogenetically activated oocytes that could develop to the blastocyst stage was higher (p data show that P4 influences the cytoplasmic maturation of porcine oocytes, at least partially, by decreasing their polyadenylation, thereby altering maternal gene expression. PMID:27672508

  19. Colicin S8 export: extracellular and cytoplasmic colicin are different.

    Science.gov (United States)

    Garcia Diaz, Maria-Elena; Concepción Curbelo, Juan Luis

    2003-12-01

    The properties of colicin S8 are different for the cytoplasmic, periplasmic and extracellular protein. Interactions with its specific receptors reflect this. Active cell extracts separate into a non-anionic along with an anionic fraction by DEAE-Sephacell chromatography. Previously, we have purified cell-associated colicin S8 as an aggregation of highly related polypeptides; cytoplasmic colicin S8 seems to be post-translationally processed into an aggregation of polypeptides of molecular mass ranging from 45,000 Da to 60,000 Da. We suggest that a conformational change to colicin S8 may occur related to the export process.

  20. Genetic expression of induced rice sterility under alien-cytoplasm

    International Nuclear Information System (INIS)

    Rice restorer lines were treated with 60Co γ-ray and 4 male sterile mutants obtained with the fertility of controlled by 4 non-allelic recessive genes, respectively. Sixty combinations were made by using male sterile plants/fertile plants as male parents, and 15 different cytoplasmic substitution lines of the same cell nucleus as female parents. The result showed that F1 spikelets were normal and fertile, and different numbers of male sterile plants were segregated in F2. Complete fertility genotype was not found among interactions between induced male sterile genes and alien-cytoplasms. (authors)

  1. Immunohistochemical study of human pulmonary surfactant apoproteins with monoclonal antibodies. Pathologic application for hyaline membrane disease.

    OpenAIRE

    Kuroki, Y.; Dempo, K.; Akino, T

    1986-01-01

    Three monoclonal antibodies, PC6, PE10, and PE12, were used for immunohistochemical studies of human lungs by immunoperoxidase staining. Monoclonal antibodies PC6 and PE10 against pulmonary surfactant apoproteins stained faint granules in the cytoplasm of some alveolar wall cells in adult lung. These stained cells appeared to be alveolar Type II cells. A fetal lung of 20 weeks' gestation had no any positive staining. However, a few scattered positive cells were observed in a newborn lung of 3...

  2. The transmission of cytoplasmic genes in Aspergillus nidulans.

    NARCIS (Netherlands)

    Coenen, A.

    1997-01-01

    IntroductionThis manuscript concerns the spread of selfish cytoplasmic genes in the fungus Aspergillus nidulans. A.nidulans is a common soil fungus that grows vegetatively by forming a network (mycelium) of hyphae and reproduces via sexual ascospores and asexual conidiospores. Cytop

  3. Optomechatronic System For Automated Intra Cytoplasmic Sperm Injection*

    Directory of Open Access Journals (Sweden)

    Shulev Assen

    2015-12-01

    Full Text Available This paper presents a complex optomechatronic system for In-Vitro Fertilization (IVF, offering almost complete automation of the Intra Cytoplasmic Sperm Injection (ICSI procedure. The compound parts and sub-systems, as well as some of the computer vision algorithms, are described below. System capabilities for ICSI have been demonstrated on infertile oocyte cells.

  4. Nuclear proteins hijacked by mammalian cytoplasmic plus strand RNA viruses

    Energy Technology Data Exchange (ETDEWEB)

    Lloyd, Richard E., E-mail: rlloyd@bcm.edu

    2015-05-15

    Plus strand RNA viruses that replicate in the cytoplasm face challenges in supporting the numerous biosynthetic functions required for replication and propagation. Most of these viruses are genetically simple and rely heavily on co-opting cellular proteins, particularly cellular RNA-binding proteins, into new roles for support of virus infection at the level of virus-specific translation, and building RNA replication complexes. In the course of infectious cycles many nuclear-cytoplasmic shuttling proteins of mostly nuclear distribution are detained in the cytoplasm by viruses and re-purposed for their own gain. Many mammalian viruses hijack a common group of the same factors. This review summarizes recent gains in our knowledge of how cytoplasmic RNA viruses use these co-opted host nuclear factors in new functional roles supporting virus translation and virus RNA replication and common themes employed between different virus groups. - Highlights: • Nuclear shuttling host proteins are commonly hijacked by RNA viruses to support replication. • A limited group of ubiquitous RNA binding proteins are commonly hijacked by a broad range of viruses. • Key virus proteins alter roles of RNA binding proteins in different stages of virus replication.

  5. Experimental Analysis of Cell Function Using Cytoplasmic Streaming

    Science.gov (United States)

    Janssens, Peter; Waldhuber, Megan

    2012-01-01

    This laboratory exercise investigates the phenomenon of cytoplasmic streaming in the fresh water alga "Nitella". Students use the fungal toxin cytochalasin D, an inhibitor of actin polymerization, to investigate the mechanism of streaming. Students use simple statistical methods to analyze their data. Typical student data are provided. (Contains 3…

  6. Nuclear proteins hijacked by mammalian cytoplasmic plus strand RNA viruses

    International Nuclear Information System (INIS)

    Plus strand RNA viruses that replicate in the cytoplasm face challenges in supporting the numerous biosynthetic functions required for replication and propagation. Most of these viruses are genetically simple and rely heavily on co-opting cellular proteins, particularly cellular RNA-binding proteins, into new roles for support of virus infection at the level of virus-specific translation, and building RNA replication complexes. In the course of infectious cycles many nuclear-cytoplasmic shuttling proteins of mostly nuclear distribution are detained in the cytoplasm by viruses and re-purposed for their own gain. Many mammalian viruses hijack a common group of the same factors. This review summarizes recent gains in our knowledge of how cytoplasmic RNA viruses use these co-opted host nuclear factors in new functional roles supporting virus translation and virus RNA replication and common themes employed between different virus groups. - Highlights: • Nuclear shuttling host proteins are commonly hijacked by RNA viruses to support replication. • A limited group of ubiquitous RNA binding proteins are commonly hijacked by a broad range of viruses. • Key virus proteins alter roles of RNA binding proteins in different stages of virus replication

  7. Method for Confirming Cytoplasmic Delivery of RNA Aptamers

    Science.gov (United States)

    Dickey, David D; Dassie, Justin P; Giangrande, Paloma H

    2016-01-01

    RNA aptamers are single-stranded RNA oligos that represent a powerful emerging technology with potential for treating numerous diseases. More recently, cell-targeted RNA aptamers have been developed for delivering RNA interference (RNAi) modulators (siRNAs and miRNAs) to specific diseased cells (e.g., cancer cells or HIV infected cells) in vitro and in vivo. However, despite initial promising reports, the broad application of this aptamer delivery technology awaits the development of methods that can verify and confirm delivery of aptamers to the cytoplasm of target cells where the RNAi machinery resides. We recently developed a functional assay (RIP assay) to confirm cellular uptake and subsequent cytoplasmic release of an RNA aptamer which binds to a cell surface receptor expressed on prostate cancer cells (PSMA). To assess cytoplasmic delivery, the aptamer was chemically conjugated to saporin, a ribosome inactivating protein toxin that is toxic to cells only when delivered to the cytoplasm (where it inhibits the ribosome) by a cell-targeting ligand (e.g., aptamer). Here, we describe the chemistry used to conjugate the aptamer to saporin and discuss a gel-based method to verify conjugation efficiency. We also detail an in vitro functional assay to confirm that the aptamer retains function following conjugation to saporin and describe a cellular assay to measure aptamer-mediated saporin-induced cytotoxicity. PMID:26472453

  8. Hepatic Subcellular Compartmentation of Cytoplasmic Phosphoenolpyruvate Carboxykinase Determined by Immunogold Electron Microscopy

    Science.gov (United States)

    Gao, Kuixiong; Cardell, Emma Lou; Morris, Randal E.; Giffin, Bruce F.; Cardell, Robert R.

    1995-08-01

    Phosphoenolpyruvate carboxykinase (PEPCK) is the rate-limiting gluconeogenic enzyme and in liver occurs in a lobular gradient from periportal to pericentral regions. The subcellular distribution of cytoplasmic PEPCK molecules within hepatocytes and its relationship to organelles have not been determined previously. In this study, we have used immunogold electron microscopy to evaluate the subcellar distribution of the enzyme, in addition to brightfield and epipolarized light microscopy. Cryosections (10 [mu]m) of perfusion-fixed rat liver were collected on silanated slides and immunostained using goat anti-rat PEPCK followed by 5-nm gold-labeled secondary and tertiary antibodies. Additionally, free-floating vibratome sections (25, 50, and 100 [mu]m) of perfusion-immersion-fixed rat liver were immunogold stained using goat anti-rat PEPCK and 5-nm gold-labeled secondary antibody, with and without silver enhancement. The immunogold labeled sections from both procedures were embedded in epoxy resin for the preparation of thin sections for electron microscopy. The results showed that the gold-labeled antibodies penetrated the entire thickness of cryosections, resulting in a high signal for PEPCK, but membranes in general, the smooth endoplasmic reticulum in particular, were not identifiable as electron dense unit membranes. On the other hand, the vibratome sections of well-fixed tissue allowed good visualization of the ultrastructure of cellular organelles, with the smooth endoplasmic reticulum appearing as vesicles and tubules with electron dense unit membranes; however, the penetration of the gold-labeled antibody was limited to cells at the surface of the vibratome sections. In both procedures, PEPCK, as indicated by gold particles, is predominantly in the glycogen areas of the cytosome and not in mitochondria, nuclei, Golgi apparatus, or other cell organelles. Hepatocytes in periportal regions have a compact subcellular distribution of PEPCK shown by gold particles

  9. Monoclonal antibodies and cancer

    International Nuclear Information System (INIS)

    The usefulness of radiolabeled monoclonal antibodies for imaging and treatment of human (ovarian) cancer was investigated. A review of tumor imaging with monoclonal antibodies is presented. Special attention is given to factors that influence the localization of the antibodies in tumors, isotope choice and methods of radiolabeling of the monoclonal antibodies. Two monoclonal antibodies, OC125 and OV-TL3, with high specificity for human epithelial ovarian cancer are characterized. A simple radio-iodination technique was developed for clinical application of the monoclonal antibodies. The behavior of monoclonal antibodies in human tumor xenograft systems and in man are described. Imaging of tumors is complicated because of high background levels of radioactivity in other sites than the tumor, especially in the bloodpool. A technique was developed to improve imaging of human tumor xenographs in nude mice, using subtraction of a specific and a non-specific antibody, radiolabeled with 111In, 67Ga and 131I. To investigate the capability of the two monoclonal antibodies, to specifically localize in human ovarian carcinomas, distribution studies in mice bearing human ovarian carcinoma xenografts were performed. One of the antibodies, OC125, was used for distribution studies in ovarian cancer patients. OC125 was used because of availability and approval to use this antibody in patients. The same antibody was used to investigate the usefulness of radioimmunoimaging in ovarian cancer patients. The interaction of injected radiolabeled antibody OC125 with circulating antigen and an assay to measure the antibody response in ovarian cancer patients after injection of the antibody is described. 265 refs.; 30 figs.; 19 tabs

  10. Phospho-Specific Antibody Probes of Intermediate Filament Proteins.

    Science.gov (United States)

    Goto, Hidemasa; Tanaka, Hiroki; Kasahara, Kousuke; Inagaki, Masaki

    2016-01-01

    Intermediate filaments (IFs) form one of the major cytoskeletal systems in the cytoplasm or beneath the nuclear membrane. Accumulating data have suggested that IF protein phosphorylation dramatically changes IF structure/dynamics in cells. For the production of an antibody recognizing site-specific protein phosphorylation (a site- and phosphorylation state-specific antibody), we first employed a strategy to immunize animals with an in vitro-phosphorylated polypeptide or a phosphopeptide (corresponding to a phosphorylated residue and its surrounding sequence of amino acids), instead of a phosphorylated protein. Our established methodology not only improves the chance of obtaining a phospho-specific antibody but also has the advantage that one can predesign a targeted phosphorylation site. It is now applied to the production of an antibody recognizing other types of site-specific posttranslational modification, such as acetylation or methylation. The use of such an antibody in immunocytochemistry enables us to analyze spatiotemporal distribution of site-specific IF protein phosphorylation. The antibody is of great use to identify a protein kinase responsible for in vivo IF protein phosphorylation and to monitor intracellular kinase activities through IF protein phosphorylation. Here, we present an overview of our methodology and describe stepwise approaches for the antibody characterization. We also provide some examples of analyses for IF protein phosphorylation involved in mitosis and signal transduction.

  11. [VGKC-complex antibodies].

    Science.gov (United States)

    Watanabe, Osamu

    2013-04-01

    Various antibodies are associated with voltage-gated potassium channels (VGKCs). Representative antibodies to VGKCs were first identified by radioimmunoassays using radioisotope-labeled alpha-dendrotoxin-VGKCs solubilized from rabbit brain. These antibodies were detected only in a proportion of patients with acquired neuromyotonia (Isaacs' syndrome). VGKC antibodies were also detected in patients with Morvan's syndrome and in those with a form of autoimmune limbic encephalitis. Recent studies indicated that the "VGKC" antibodies are mainly directed toward associated proteins (for example LGI-1 and CASPR-2) that complex with the VGKCs themselves. The "VGKC" antibodies are now commonly known as VGKC-complex antibodies. In general, LGI-1 antibodies are most commonly detected in patients with limbic encephalitis with syndrome of inappropriate secretion of antidiuretic hormone. CASPR-2 antibodies are present in the majority of patients with Morvan's syndrome. These patients develop combinations of CNS symptoms, autonomic dysfunction, and peripheral nerve hyperexcitability. Furthermore, VGKC-complex antibodies are tightly associated with chronic idiopathic pain. Hyperexcitability of nociceptive pathways has also been implicated. These antibodies may be detected in sera of some patients with neurodegenerative diseases (for example, amyotrophic lateral sclerosis and Creutzfeldt-Jakob disease).

  12. Bartonella henselae endocarditis in an immunocompetent adult.

    Science.gov (United States)

    Holmes, A H; Greenough, T C; Balady, G J; Regnery, R L; Anderson, B E; O'Keane, J C; Fonger, J D; McCrone, E L

    1995-10-01

    We describe a case of aggressive Bartonella henselae endocarditis in an immunocompetent man who owned a cat. Aortic valve replacement was required, and his infection was diagnosed by histology, serology, and polymerase chain reaction analysis. The manifestations of his disease included mediastinal lymphadenopathy, glomerulonephritis, myocarditis, and a petechial rash; the unusual finding of a positive titer of c-antineutrophil cytoplasmic antibodies was noted. Serological titers were markedly elevated for > 1 year despite clinical improvement. PMID:8645787

  13. Eosinophilic Granulomatosis with Polyangiitis: An Overview

    OpenAIRE

    Gioffredi, Andrea; Maritati, Federica; Oliva, Elena; Buzio, Carlo

    2014-01-01

    Eosinophilic granulomatosis with polyangiitis (EGPA) is a multisystemic disorder, belonging to the small vessel anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis, defined as an eosinophil-rich and necrotizing granulomatous inflammation often involving the respiratory tract, and necrotizing vasculitis predominantly affecting small to medium-sized vessels, associated with asthma and eosinophilia. EGPA pathogenesis is not well known: HLA-DRB1*04 and *07, HLA-DRB4 and IL10.2 haplo...

  14. “Peripheral Neuropathy Crippling Bronchial Asthma”: Two Rare Case Reports of Churg-Strauss Syndrome

    Directory of Open Access Journals (Sweden)

    Kamal Kishore Pandita

    2014-01-01

    Full Text Available Churg-Strauss syndrome (CSS is a rare cause of vasculitic neuropathy. Although rare and potentially fatal, Churg-Strauss syndrome (CSS is easily diagnosable and treatable. The presence of bronchial asthma with peripheral neuropathy in a patient alerts a physician to this diagnosis. This is vividly illustrated by the presented two cases who had neuropathy associated with bronchial asthma, eosinophilia, sinusitis, and positive perinuclear antineutrophil cytoplasmic antibodies (p-ANCA test, which improved with administration of steroids.

  15. Serum levels of KL-6 reflect disease activity of interstitial pneumonia associated with ANCA-related vasculitis

    OpenAIRE

    Iwata, Yasunori; Wada, Takashi; Furuichi, Kengo; Kitagawa, Kiyoki; Kokubo, Satoshi; Kobayashi, Motoo; Sakai, Norihiko; Yoshimoto, Keiichi; Shimizu, Miho; Kobayashi, Kenichi; Yokoyama, Hitoshi

    2001-01-01

    Objective. KL-6 is reported to be excreted from the lung alveolar and bronchial epithelial cells and may be a good marker for monitoring disease activity of interstitial pneumonia. This study was designed to ascertain the clinical significance of serum KL-6 levels in interstitial pneumonia associated with anti-neutrophil cytoplasmic antibody (ANCA)-related vasculitis. Methods. Serum KL-6 levels were determined by an enzyme-linked immunosorbent assay. Patients. We examined 20 healthy subjects,...

  16. Microscopic polyangiitis complicated with ileal involvement detected by double-balloon endoscopy: a case report

    OpenAIRE

    Fukushima, Masashi; Inoue, Satoko; Ono, Yuichiro; Tamaki, Yoshitaka; Yoshimura, Hajime; Imai, Yukihiro; Inokuma, Tetsuro

    2013-01-01

    Background Microscopic polyangiitis is characterized by pauci-immune, necrotizing small-vessel vasculitis and an anti-neutrophil cytoplasmic antibody-associated vasculitis. Although gastrointestinal involvement in microscopic polyangiitis is not rare, endoscopic observation of it is extremely rare. To the best of our knowledge, this is the first case report of small intestinal involvement in microscopic polyangiitis detected and followed up by double-balloon endoscopy. Case presentation A 70-...

  17. The role of conjunctival biopsy in the diagnosis of granulomatosis with polyangiitis

    OpenAIRE

    Ursea, Roxana; De Castro, Dawn; Bowen, Trent J.; Chan, Chi-Chao

    2015-01-01

    Background The purpose of this study is to describe a patient who was diagnosed with granulomatosis with polyangiitis based on conjunctival biopsy. This study is a case report and review of the literature. Findings A 48-year-old Caucasian woman presented with a 2-week history of a left eye peripheral corneal ulcer with adjacent conjunctivitis and a 4-month history of a non-resolving productive cough. Given her elevated serum perinuclear antineutrophil cytoplasmic antibody (P-ANCA) and erythro...

  18. Drug-Induced Bullous Sweet Syndrome with Multiple Autoimmune Features

    OpenAIRE

    2010-01-01

    Sweet syndrome (SS) (Acute Febrile Neutrophilic Dermatosis) has been reported in association with autoimmune phenomena including relapsing polychondritis, drug-induced lupus, and the development of antineutrophil cytoplasmic antibodies (ANCAs). However, a combination of these autoimmune features has not been reported. Herein, we report a case of drug-induced bullous SS with ocular and mucosal involvement, glomerulonephritis, and multiple autoimmune features including clinical polychondritis w...

  19. Hypertrophic cranial pachymeningitis in mpo-anca-related vasculitis: a case report and literature review

    OpenAIRE

    Watanabe, Kimio; Tani, Yoshihiro; Kimura, Hiroshi; Asai, Jun; Tanaka, Kenichi; Hayashi, Yoshimitsu; Asahi, Koichi; Nakayama, Masaaki; Watanabe, Tsuyoshi

    2013-01-01

    A 75-year-old woman presented with rapidly progressive glomerulonephritis with positive results for anti-myeloperoxidase anti-neutrophil cytoplasmic antibody (MPO-ANCA). Corticosteroid therapy was successfully introduced. However, 7 months later, magnetic resonance imaging revealed marked swelling in the falx cerebri and high density regions were apparent on gallium scintigraphy, leading to diagnosis of hypertrophic cranial pachymeningitis (HCP). Symptoms improved with intensified corticoster...

  20. BPI-ANCA Provides Additional Clinical Information to Anti-Pseudomonas Serology: Results from a Cohort of 117 Swedish Cystic Fibrosis Patients

    OpenAIRE

    Ulrika Lindberg; Malin Carlsson; Thomas Hellmark; Mårten Segelmark

    2015-01-01

    Patients with cystic fibrosis (CF) colonized with Pseudomonas aeruginosa (P. aeruginosa) have worse prognosis compared with patients who are not. BPI-ANCA is an anti-neutrophil cytoplasmic antibody against BPI (bactericidal/permeability increasing protein) correlating with P. aeruginosa colonization and adverse long time prognosis. Whether it provides additional information as compared to standard anti-P. aeruginosa serology tests is not known. 117 nontransplanted CF patients at the CF centre...

  1. Histone H2B-IFI16 Recognition of Nuclear Herpesviral Genome Induces Cytoplasmic Interferon-β Responses

    Science.gov (United States)

    Iqbal, Jawed; Ansari, Mairaj Ahmed; Kumar, Binod; Dutta, Dipanjan; Roy, Arunava; Chikoti, Leela; Pisano, Gina; Dutta, Sujoy; Veettil, Mohanan Valiya; Chandran, Bala

    2016-01-01

    IFI16 (gamma-interferon-inducible protein 16), a predominantly nuclear protein involved in transcriptional regulation, also functions as an innate immune response DNA sensor and induces the IL-1β and antiviral type-1 interferon-β (IFN-β) cytokines. We have shown that IFI16, in association with BRCA1, functions as a sequence independent nuclear sensor of episomal dsDNA genomes of KSHV, EBV and HSV-1. Recognition of these herpesvirus genomes resulted in IFI16 acetylation, BRCA1-IFI16-ASC-procaspase-1 inflammasome formation, cytoplasmic translocation, and IL-1β generation. Acetylated IFI16 also interacted with cytoplasmic STING and induced IFN-β. However, the identity of IFI16 associated nuclear proteins involved in STING activation and the mechanism is not known. Mass spectrometry of proteins precipitated by anti-IFI16 antibodies from uninfected endothelial cell nuclear lysate revealed that histone H2B interacts with IFI16. Single and double proximity ligation microscopy, immunoprecipitation, EdU-genome labeled virus infection, and chromatin immunoprecipitation studies demonstrated that H2B is associated with IFI16 and BRCA1 in the nucleus in physiological conditions. De novo KSHV and HSV-1 infection as well as latent KSHV and EBV infection induces the cytoplasmic distribution of H2B-IFI16, H2B-BRCA1 and IFI16-ASC complexes. Vaccinia virus (dsDNA) cytoplasmic replication didn’t induce the redistribution of nuclear H2B-IFI16 or H2B into the cytoplasm. H2B is critical in KSHV and HSV-1 genome recognition by IFI16 during de novo infection. Viral genome sensing by IFI16-H2B-BRCA1 leads to BRCA1 dependent recruitment of p300, and acetylation of H2B and IFI16. BRCA1 knockdown or inhibition of p300 abrogated the acetylation of H2B-IFI16 or H2B. Ran-GTP protein mediated the translocation of acetylated H2B and IFI16 to the cytoplasm along with BRCA1 that is independent of IFI16-ASC inflammasome. ASC knockdown didn’t affect the acetylation of H2B, its cytoplasmic

  2. Cytoplasmic genetic variation and extensive cytonuclear interactions influence natural variation in the metabolome

    DEFF Research Database (Denmark)

    Joseph, Bindu; Corwin, Jason A.; Li, Baohua;

    2013-01-01

    affects phenotypic variation. This showed that the cytoplasmic variation had effects similar to, if not larger than, the largest individual nuclear locus. Inclusion of cytoplasmic variation into the genetic model greatly increased the explained phenotypic variation. Cytoplasmic genetic variation was a...... central hub in the epistatic network controlling the plant metabolome. This epistatic influence manifested such that the cytoplasmic background could alter or hide pairwise epistasis between nuclear loci. Thus, cytoplasmic genetic variation plays a central role in controlling natural variation in...... metabolomic networks. This suggests that cytoplasmic genomes must be included in any future analysis of natural variation....

  3. Mapping of ESE-1 subdomains required to initiate mammary epithelial cell transformation via a cytoplasmic mechanism

    Directory of Open Access Journals (Sweden)

    Tentler John J

    2011-08-01

    Full Text Available Abstract Background The ETS family transcription factor ESE-1 is often overexpressed in human breast cancer. ESE-1 initiates transformation of MCF-12A cells via a non-transcriptional, cytoplasmic process that is mediated by a unique 40-amino acid serine and aspartic acid rich (SAR subdomain, whereas, ESE-1's nuclear transcriptional property is required to maintain the transformed phenotype of MCF7, ZR-75-1 and T47D breast cancer cells. Results To map the minimal functional nuclear localization (NLS and nuclear export (NES signals, we fused in-frame putative NLS and NES motifs between GFP and the SAR domain. Using these GFP constructs as reporters of subcellular localization, we mapped a single NLS to six basic amino acids (242HGKRRR247 in the AT-hook and two CRM1-dependent NES motifs, one to the pointed domain (NES1: 102LCNCALEELRL112 and another to the DNA binding domain (DBD, (NES2: 275LWEFIRDILI284. Moreover, analysis of a putative NLS located in the DBD (316GQKKKNSN323 by a similar GFP-SAR reporter or by internal deletion of the DBD, revealed this sequence to lack NLS activity. To assess the role of NES2 in regulating ESE-1 subcellular localization and subsequent transformation potency, we site-specifically mutagenized NES2, within full-length GFP-ESE-1 and GFP-NES2-SAR reporter constructs. These studies show that site-specific mutation of NES2 completely abrogates ESE-1 transforming activity. Furthermore, we show that exclusive cytoplasmic targeting of the SAR domain is sufficient to initiate transformation, and we report that an intact SAR domain is required, since block mutagenesis reveals that an intact SAR domain is necessary to maintain its full transforming potency. Finally, using a monoclonal antibody targeting the SAR domain, we demonstrate that the SAR domain contains a region accessible for protein - protein interactions. Conclusions These data highlight that ESE-1 contains NLS and NES signals that play a critical role in

  4. Cytoplasmic streaming in plant cells: the role of wall slip.

    Science.gov (United States)

    Wolff, K; Marenduzzo, D; Cates, M E

    2012-06-01

    We present a computer simulation study, via lattice Boltzmann simulations, of a microscopic model for cytoplasmic streaming in algal cells such as those of Chara corallina. We modelled myosin motors tracking along actin lanes as spheres undergoing directed motion along fixed lines. The sphere dimension takes into account the fact that motors drag vesicles or other organelles, and, unlike previous work, we model the boundary close to which the motors move as walls with a finite slip layer. By using realistic parameter values for actin lane and myosin density, as well as for endoplasmic and vacuole viscosity and the slip layer close to the wall, we find that this simplified view, which does not rely on any coupling between motors, cytoplasm and vacuole other than that provided by viscous Stokes flow, is enough to account for the observed magnitude of streaming velocities in intracellular fluid in living plant cells.

  5. Genetic analysis of the cytoplasmic dynein subunit families.

    Directory of Open Access Journals (Sweden)

    K Kevin Pfister

    2006-01-01

    Full Text Available Cytoplasmic dyneins, the principal microtubule minus-end-directed motor proteins of the cell, are involved in many essential cellular processes. The major form of this enzyme is a complex of at least six protein subunits, and in mammals all but one of the subunits are encoded by at least two genes. Here we review current knowledge concerning the subunits, their interactions, and their functional roles as derived from biochemical and genetic analyses. We also carried out extensive database searches to look for new genes and to clarify anomalies in the databases. Our analysis documents evolutionary relationships among the dynein subunits of mammals and other model organisms, and sheds new light on the role of this diverse group of proteins, highlighting the existence of two cytoplasmic dynein complexes with distinct cellular roles.

  6. Nucleoporin Nup98 mediates galectin-3 nuclear-cytoplasmic trafficking

    International Nuclear Information System (INIS)

    Highlights: •Nuclear pore protein Nup98 is a novel binding partner of galectin-3. •Nup98 transports galectin-3 into cytoplasm. •Nup98 depletion leads to galectin-3 nuclear transport and induces growth retardation. •Nup98 may involve in ß-catenin pathway through interaction with galectin-3. -- Abstract: Nucleoporin Nup98 is a component of the nuclear pore complex, and is important in transport across the nuclear pore. Many studies implicate nucleoporin in cancer progression, but no direct mechanistic studies of its effect in cancer have been reported. We show here that Nup98 specifically regulates nucleus–cytoplasm transport of galectin-3, which is a ß-galactoside-binding protein that affects adhesion, migration, and cancer progression, and controls cell growth through the ß-catenin signaling pathway in cancer cells. Nup98 interacted with galectin-3 on the nuclear membrane, and promoted galectin-3 cytoplasmic translocation whereas other nucleoporins did not show these functions. Inversely, silencing of Nup98 expression by siRNA technique localized galectin-3 to the nucleus and retarded cell growth, which was rescued by Nup98 transfection. In addition, Nup98 RNA interference significantly suppressed downstream mRNA expression in the ß-catenin pathway, such as cyclin D1 and FRA-1, while nuclear galectin-3 binds to ß-catenin to inhibit transcriptional activity. Reduced expression of ß-catenin target genes is consistent with the Nup98 reduction and the galectin-3–nucleus translocation rate. Overall, the results show Nup98’s involvement in nuclear–cytoplasm translocation of galectin-3 and ß-catenin signaling pathway in regulating cell proliferation, and the results depicted here suggest a novel therapeutic target/modality for cancers

  7. Influenza a virus assembly intermediates fuse in the cytoplasm.

    Science.gov (United States)

    Lakdawala, Seema S; Wu, Yicong; Wawrzusin, Peter; Kabat, Juraj; Broadbent, Andrew J; Lamirande, Elaine W; Fodor, Ervin; Altan-Bonnet, Nihal; Shroff, Hari; Subbarao, Kanta

    2014-03-01

    Reassortment of influenza viral RNA (vRNA) segments in co-infected cells can lead to the emergence of viruses with pandemic potential. Replication of influenza vRNA occurs in the nucleus of infected cells, while progeny virions bud from the plasma membrane. However, the intracellular mechanics of vRNA assembly into progeny virions is not well understood. Here we used recent advances in microscopy to explore vRNA assembly and transport during a productive infection. We visualized four distinct vRNA segments within a single cell using fluorescent in situ hybridization (FISH) and observed that foci containing more than one vRNA segment were found at the external nuclear periphery, suggesting that vRNA segments are not exported to the cytoplasm individually. Although many cytoplasmic foci contain multiple vRNA segments, not all vRNA species are present in every focus, indicating that assembly of all eight vRNA segments does not occur prior to export from the nucleus. To extend the observations made in fixed cells, we used a virus that encodes GFP fused to the viral polymerase acidic (PA) protein (WSN PA-GFP) to explore the dynamics of vRNA assembly in live cells during a productive infection. Since WSN PA-GFP colocalizes with viral nucleoprotein and influenza vRNA segments, we used it as a surrogate for visualizing vRNA transport in 3D and at high speed by inverted selective-plane illumination microscopy. We observed cytoplasmic PA-GFP foci colocalizing and traveling together en route to the plasma membrane. Our data strongly support a model in which vRNA segments are exported from the nucleus as complexes that assemble en route to the plasma membrane through dynamic colocalization events in the cytoplasm. PMID:24603687

  8. CD248 facilitates tumor growth via its cytoplasmic domain

    International Nuclear Information System (INIS)

    Stromal fibroblasts participate in the development of a permissive environment for tumor growth, yet molecular pathways to therapeutically target fibroblasts are poorly defined. CD248, also known as endosialin or tumor endothelial marker 1 (TEM1), is a transmembrane glycoprotein expressed on activated fibroblasts. We recently showed that the cytoplasmic domain of CD248 is important in facilitating an inflammatory response in a mouse model of arthritis. Others have reported that CD248 gene inactivation in mice results in dampened tumor growth. We hypothesized that the conserved cytoplasmic domain of CD248 is important in regulating tumor growth. Mice lacking the cytoplasmic domain of CD248 (CD248CyD/CyD) were generated and evaluated in tumor models, comparing the findings with wild-type mice (CD248WT/WT). As compared to the response in CD248WT/WT mice, growth of T241 fibrosarcomas and Lewis lung carcinomas was significantly reduced in CD248CyD/CyD mice. Tumor size was similar to that seen with CD248-deficient mice. Conditioned media from CD248CyD/CyD fibroblasts were less effective at supporting T241 fibrosarcoma cell survival. In addition to our previous observation of reduced release of activated matrix metalloproteinase (MMP)-9, CD248CyD/CyD fibroblasts also had impaired PDGF-BB-induced migration and expressed higher transcripts of tumor suppressor factors, transgelin (SM22α), Hes and Hey1. The multiple pathways regulated by the cytoplasmic domain of CD248 highlight its potential as a therapeutic target to treat cancer

  9. CD248 facilitates tumor growth via its cytoplasmic domain

    Directory of Open Access Journals (Sweden)

    Janssens Tom

    2011-05-01

    Full Text Available Abstract Background Stromal fibroblasts participate in the development of a permissive environment for tumor growth, yet molecular pathways to therapeutically target fibroblasts are poorly defined. CD248, also known as endosialin or tumor endothelial marker 1 (TEM1, is a transmembrane glycoprotein expressed on activated fibroblasts. We recently showed that the cytoplasmic domain of CD248 is important in facilitating an inflammatory response in a mouse model of arthritis. Others have reported that CD248 gene inactivation in mice results in dampened tumor growth. We hypothesized that the conserved cytoplasmic domain of CD248 is important in regulating tumor growth. Methods Mice lacking the cytoplasmic domain of CD248 (CD248CyD/CyD were generated and evaluated in tumor models, comparing the findings with wild-type mice (CD248WT/WT. Results As compared to the response in CD248WT/WT mice, growth of T241 fibrosarcomas and Lewis lung carcinomas was significantly reduced in CD248CyD/CyD mice. Tumor size was similar to that seen with CD248-deficient mice. Conditioned media from CD248CyD/CyD fibroblasts were less effective at supporting T241 fibrosarcoma cell survival. In addition to our previous observation of reduced release of activated matrix metalloproteinase (MMP-9, CD248CyD/CyD fibroblasts also had impaired PDGF-BB-induced migration and expressed higher transcripts of tumor suppressor factors, transgelin (SM22α, Hes and Hey1. Conclusions The multiple pathways regulated by the cytoplasmic domain of CD248 highlight its potential as a therapeutic target to treat cancer.

  10. Influenza a virus assembly intermediates fuse in the cytoplasm.

    Directory of Open Access Journals (Sweden)

    Seema S Lakdawala

    2014-03-01

    Full Text Available Reassortment of influenza viral RNA (vRNA segments in co-infected cells can lead to the emergence of viruses with pandemic potential. Replication of influenza vRNA occurs in the nucleus of infected cells, while progeny virions bud from the plasma membrane. However, the intracellular mechanics of vRNA assembly into progeny virions is not well understood. Here we used recent advances in microscopy to explore vRNA assembly and transport during a productive infection. We visualized four distinct vRNA segments within a single cell using fluorescent in situ hybridization (FISH and observed that foci containing more than one vRNA segment were found at the external nuclear periphery, suggesting that vRNA segments are not exported to the cytoplasm individually. Although many cytoplasmic foci contain multiple vRNA segments, not all vRNA species are present in every focus, indicating that assembly of all eight vRNA segments does not occur prior to export from the nucleus. To extend the observations made in fixed cells, we used a virus that encodes GFP fused to the viral polymerase acidic (PA protein (WSN PA-GFP to explore the dynamics of vRNA assembly in live cells during a productive infection. Since WSN PA-GFP colocalizes with viral nucleoprotein and influenza vRNA segments, we used it as a surrogate for visualizing vRNA transport in 3D and at high speed by inverted selective-plane illumination microscopy. We observed cytoplasmic PA-GFP foci colocalizing and traveling together en route to the plasma membrane. Our data strongly support a model in which vRNA segments are exported from the nucleus as complexes that assemble en route to the plasma membrane through dynamic colocalization events in the cytoplasm.

  11. Nucleoporin Nup98 mediates galectin-3 nuclear-cytoplasmic trafficking

    Energy Technology Data Exchange (ETDEWEB)

    Funasaka, Tatsuyoshi, E-mail: funasaka@staff.kanazawa-u.ac.jp [Laboratory of Molecular and Cellular Biology, Department of Biology, Faculty of Natural Systems, Institute of Science and Engineering, Kanazawa University, Ishikawa (Japan); Balan, Vitaly; Raz, Avraham [Department of Oncology and Pathology, Karmanos Cancer Institute, Wayne State University, School of Medicine, Detroit, MI (United States); Wong, Richard W., E-mail: rwong@staff.kanazawa-u.ac.jp [Laboratory of Molecular and Cellular Biology, Department of Biology, Faculty of Natural Systems, Institute of Science and Engineering, Kanazawa University, Ishikawa (Japan); Bio-AFM Frontier Research Center, Kanazawa Kanazawa University, Ishikawa (Japan)

    2013-04-26

    Highlights: •Nuclear pore protein Nup98 is a novel binding partner of galectin-3. •Nup98 transports galectin-3 into cytoplasm. •Nup98 depletion leads to galectin-3 nuclear transport and induces growth retardation. •Nup98 may involve in ß-catenin pathway through interaction with galectin-3. -- Abstract: Nucleoporin Nup98 is a component of the nuclear pore complex, and is important in transport across the nuclear pore. Many studies implicate nucleoporin in cancer progression, but no direct mechanistic studies of its effect in cancer have been reported. We show here that Nup98 specifically regulates nucleus–cytoplasm transport of galectin-3, which is a ß-galactoside-binding protein that affects adhesion, migration, and cancer progression, and controls cell growth through the ß-catenin signaling pathway in cancer cells. Nup98 interacted with galectin-3 on the nuclear membrane, and promoted galectin-3 cytoplasmic translocation whereas other nucleoporins did not show these functions. Inversely, silencing of Nup98 expression by siRNA technique localized galectin-3 to the nucleus and retarded cell growth, which was rescued by Nup98 transfection. In addition, Nup98 RNA interference significantly suppressed downstream mRNA expression in the ß-catenin pathway, such as cyclin D1 and FRA-1, while nuclear galectin-3 binds to ß-catenin to inhibit transcriptional activity. Reduced expression of ß-catenin target genes is consistent with the Nup98 reduction and the galectin-3–nucleus translocation rate. Overall, the results show Nup98’s involvement in nuclear–cytoplasm translocation of galectin-3 and ß-catenin signaling pathway in regulating cell proliferation, and the results depicted here suggest a novel therapeutic target/modality for cancers.

  12. Heavy chain only antibodies

    DEFF Research Database (Denmark)

    Moghimi, Seyed Moein; Rahbarizadeh, Fatemeh; Ahmadvand, Davoud;

    2013-01-01

    Unlike conventional antibodies, heavy chain only antibodies derived from camel contain a single variable domain (VHH) and two constant domains (CH2 and CH3). Cloned and isolated VHHs possess unique properties that enable them to excel conventional therapeutic antibodies and their smaller antigen-...... for combating HER2+ breast cancer. © 2013 by Tabriz University of Medical Sciences.......Unlike conventional antibodies, heavy chain only antibodies derived from camel contain a single variable domain (VHH) and two constant domains (CH2 and CH3). Cloned and isolated VHHs possess unique properties that enable them to excel conventional therapeutic antibodies and their smaller antigen......-binding fragments in cancer targeting and therapy. VHHs express low immunogenicity, are highly robust and easy to manufacture and have the ability to recognize hidden or uncommon epitopes. We highlight the utility of VHH in design of new molecular, multifunctional particulate and immune cell-based systems...

  13. Engineering antibody therapeutics.

    Science.gov (United States)

    Chiu, Mark L; Gilliland, Gary L

    2016-06-01

    The successful introduction of antibody-based protein therapeutics into the arsenal of treatments for patients has within a few decades fostered intense innovation in the production and engineering of antibodies. Reviewed here are the methods currently used to produce antibodies along with how our knowledge of the structural and functional characterization of immunoglobulins has resulted in the engineering of antibodies to produce protein therapeutics with unique properties, both biological and biophysical, that are leading to novel therapeutic approaches. Antibody engineering includes the introduction of the antibody combining site (variable regions) into a host of architectures including bi and multi-specific formats that further impact the therapeutic properties leading to further advantages and successes in patient treatment. PMID:27525816

  14. THE KINETICS OF CYTOPLASMIC GRANULE SECRETION IN NATURAL KILLER CYTOTOXICITY

    Institute of Scientific and Technical Information of China (English)

    龚伊红; R.R.Hcrberman; C.W.Reynolds

    1994-01-01

    Antisexum against purified cytoplasmic granules from rat LGL tumor cells, and protein A-gold inmmnoelec-tron microscopy were used to study the secretory events in lysis of YAC-1 tumor cells by rat LGL tumor cells or by isolated LGL from normal rats. After 30 min incubation of effector and target cells together, gold-labeled cyto-plasmic granules were often seen concentrated in the area of the LGL adjacent to the ~ YAC-1 Within 60min,the grantees were observed to move to the cell border near the conjugazed site. At this point, fine granules were fused with file cell membrane, and subsequently released file gold-labeled contents into the junction between the LGL and the target cell. Gold particles could be seen at the B-T interface, on the surface,or sometimes on the target cell surface.These data provide direct evidence for the hypothesis that under conditions of active cytotoxicity,natural killer cells secrete their cytoplasmic granule contents leading to the deposition of granule material on the target cell surface and the eventual lysis of the cell.

  15. Sequences determining the cytoplasmic localization of a chemoreceptor domain.

    Science.gov (United States)

    Seligman, L; Bailey, J; Manoil, C

    1995-01-01

    The Escherichia coli serine chemoreceptor (Tsr) is a protein with a simple topology consisting of two membrane-spanning sequences (TM1 and TM2) separating a large periplasmic domain from N-terminal and C-terminal cytoplasmic regions. We analyzed the contributions of several sequence elements to the cytoplasmic localization of the C-terminal domain by using chemoreceptor-alkaline phosphatase gene fusions. The principal findings were as follows. (i) The cytoplasmic localization of the C-terminal domain depended on TM2 but was quite tolerant of mutations partially deleting or introducing charged residues into the sequence. (ii) The basal level of C-terminal domain export was significantly higher in proteins with the wild-type periplasmic domain than in derivatives with a shortened periplasmic domain, suggesting that the large size of the wild-type domain promotes partial membrane misinsertion. (iii) The membrane insertion of deletion derivatives with a single spanning segment (TM1 or TM2) could be controlled by either an adjacent positively charged sequence or an adjacent amphipathic sequence. The results provide evidence that the generation of the Tsr membrane topology is an overdetermined process directed by an interplay of sequences promoting and opposing establishment of the normal structure. PMID:7730259

  16. Ionizing Radiation Induces HMGB1 Cytoplasmic Translocation and Extracellular Release

    Institute of Scientific and Technical Information of China (English)

    Lili Wang; Li He; Guoqiang Bao; Xin He; Saijun Fan; Haichao Wang

    2016-01-01

    Objective A nucleosomal protein,HMGBI,can be secreted by activated immune cells or passively released by dying cells,thereby amplifying rigorous inflammatory responses.In this study we aimed to test the possibility that radiation similarly induces cytoplasmic HMGB1 translocation and release.Methods Human skin fibroblast (GM0639) and bronchial epithelial (16HBE) cells and rats were exposed to X-ray radiation,and HMGB1 translocation and release were then assessed by immunocytochemistry and immunoassay,respectively.Results At a wide dose range(4.0-12.0 Gy),X-ray radiation induced a dramatic cytoplasmic HMGB1 translocation,and triggered a time-and dose-dependent HMGB1 release both in vitro and in vivo.The radiation-mediated HMGB1 release was also associated with noticeable chromosomal DNA damage and loss of cell viability.Conclusions Radiation induces HMGB1 cytoplasmic translocation and extracellular release through active secretion and passive leakage processes.

  17. Infection of human and non-human cells by a highly fusogenic primary CD4-independent HIV-1 isolate with a truncated envelope cytoplasmic tail

    International Nuclear Information System (INIS)

    Truncation of the envelope cytoplasmic tail has enabled FIV, SIV, and some laboratory HIV-1 strains to acquire broader cellular tropism and enhanced fusogenicity. Here we have characterized a primary CD4-independent HIV-1 isolate (92UG046-T8) with a truncated cytoplasmic tail that was able to infect and induce syncytia in primary lymphocytes from human, chimpanzee, and monkey, as well as CD4-negative cell lines from human and monkey. Increased syncytia were also noticeable with 293 cells expressing the cloned envelope from the 92UG046-T8 isolate suggesting envelope-mediated cellular fusion. Except pooled serum from HIV-1-infected individuals, monoclonal anti-envelope antibodies or antibodies/antagonists against CD4, CXCR4, and CCR5 were not able to prevent infection by the 92UG046-T8 isolate. This is the first report showing a primary HIV-1 variant with truncated cytoplasmic tail which is highly fusogenic and can infect a broad range of cells from human and non-human origins. In vivo evolution of similar HIV-1 mutants may have important implications in AIDS pathogenesis

  18. Reactivity of monoclonal antibodies to species-specific antigens of Entamoeba histolytica.

    Science.gov (United States)

    Tachibana, H; Kobayashi, S; Nagakura, K; Kaneda, Y; Takeuchi, T

    1991-01-01

    Twenty monoclonal antibodies were produced against trophozoites of Entamoeba histolytica strains HK-9 and HM-1: IMSS. When reactivity to various enteric protozoa was examined by an indirect fluorescence antibody test, 15 of the monoclonal antibodies were strongly reactive with E. histolytica trophozoites. Species-specific antigens recognized by these monoclonal antibodies were located on the plasma membrane, nucleus, cytoplasm, and cytoskeletal structures of the trophozoites. Two of the remaining five monoclonals reacted strongly with trophozoites of the E. histolytica-like Laredo strain. The determinant antigen was located in the cytoplasm. The three remaining monoclonal antibodies were found to recognize cross-reactive antigens between E. histolytica and E. histolytica-like Laredo, E. hartmanni, E. coli, Dientamoeba fragilis, Giardia lamblia, and Trichomonas hominis. These three antibodies were also reactive with T. vaginalis and mammalian cells such as HeLa cells. Thus, the combined use of monoclonal antibodies seems capable of distinguishing E. histolytica and/or E. histolytica-like Laredo from other enteric protozoa. PMID:1724012

  19. Requirements for, and Cytoplasmic Concentrations of, Sulphate and Chloride, and Cytoplasmic Volume Spaces in the Halophilic Bacterium Ectothiorhodospira mobilis

    OpenAIRE

    Imhoff, Johannes F.; Riedel, Thomas

    1989-01-01

    Ectothiorhodospira mobilis is a halophilic phototrophic bacterium that has been isolated from soda lakes containing high concentrations of sulphate, chloride and carbonates. It utilizes reduced sulphur compounds as photosynthetic electron donors and oxidizes them to sulphate, but can also grow photoheterotrophically with sulphate as sole sulphur source. The requirements for, and the cytoplasmic concentrations of, sulphate and chloride have been determined. High concentrations of sulphate are ...

  20. A PSTAIRE CDK-like protein localizes in nuclei and cytoplasm of Physarum polycephalum and functions in the mitosis

    Institute of Scientific and Technical Information of China (English)

    2004-01-01

    CDKs play key roles in controlling cell cycle progression in all eukaryotes. In plants, multiple CDKs are present,among which the best characterized CDKs are PSTAIRE CDKs. In this study, we carried out Western blot,immunoelectron microscopy and antibody treatment with an anti-PSTAIRE monoclonal antibody to explore the subcellular localization and functions of PSTAIRE CDKs in Physarum polycephalum. The results of Western blot and immunoelectron microscopy showed that in P. polycephalum, a PSTAIRE CDK-like protein was 34 kD in molecular weight and located in both nuclei and cytoplasm. In nuclei, the protein was mainly associated with chromosomes and nucleoli. The expression of the PSTAIRE CDK-like protein in both the plasmodia and nuclei showed little fluctuation through the whole cell cycle. When treated with an anti-PSTAIRE monoclonal antibody at early S phase, the cells were arrested in S phase, and the mitotic onset of P. polycephalum was blocked for about 1 h when treated at early G2 phase.Our data indicated that the PSTAIRE CDK- like protein has a direct bearing on the mitosis.

  1. DMPD: Negative regulation of cytoplasmic RNA-mediated antiviral signaling. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 18703349 Negative regulation of cytoplasmic RNA-mediated antiviral signaling. Komur...Show Negative regulation of cytoplasmic RNA-mediated antiviral signaling. PubmedID 18703349 Title Negative r...egulation of cytoplasmic RNA-mediated antiviral signaling. Authors Komuro A, Bamm

  2. RBC Antibody Screen

    Science.gov (United States)

    ... the baby is Rh-positive and the mother's antibody status is negative for anti-D, the mother is given additional RhIG. This test also may be used to help diagnose autoimmune-related hemolytic anemia ... when a person produces antibodies against his or her own RBC antigens. This ...

  3. Selection of Recombinant Human Antibodies.

    Science.gov (United States)

    Tomszak, Florian; Weber, Susanne; Zantow, Jonas; Schirrmann, Thomas; Hust, Michael; Frenzel, André

    2016-01-01

    Since the development of therapeutic antibodies the demand of recombinant human antibodies is steadily increasing. Traditionally, therapeutic antibodies were generated by immunization of rat or mice, the generation of hybridoma clones, cloning of the antibody genes and subsequent humanization and engineering of the lead candidates. In the last few years, techniques were developed that use transgenic animals with a human antibody gene repertoire. Here, modern recombinant DNA technologies can be combined with well established immunization and hybridoma technologies to generate already affinity maturated human antibodies. An alternative are in vitro technologies which enabled the generation of fully human antibodies from antibody gene libraries that even exceed the human antibody repertoire. Specific antibodies can be isolated from these libraries in a very short time and therefore reduce the development time of an antibody drug at a very early stage.In this review, we describe different technologies that are currently used for the in vitro and in vivo generation of human antibodies. PMID:27236551

  4. Anti-insulin antibody test

    Science.gov (United States)

    Insulin antibodies - serum; Insulin Ab test; Insulin resistance - insulin antibodies; Diabetes - insulin antibodies ... You appear to have an allergic response to insulin Insulin no longer seems to control your diabetes

  5. Cytoplasmic TRADD Confers a Worse Prognosis in Glioblastoma

    Directory of Open Access Journals (Sweden)

    Sharmistha Chakraborty

    2013-08-01

    Full Text Available Tumor necrosis factor receptor 1 (TNFR1-associated death domain protein (TRADD is an important adaptor in TNFR1 signaling and has an essential role in nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB activation and survival signaling. Increased expression of TRADD is sufficient to activate NF-κB. Recent studies have highlighted the importance of NF-κB activation as a key pathogenic mechanism in glioblastoma multiforme (GBM, the most common primary malignant brain tumor in adults.We examined the expression of TRADD by immunohistochemistry (IHC and find that TRADD is commonly expressed at high levels in GBM and is detected in both cytoplasmic and nuclear distribution. Cytoplasmic IHC TRADD scoring is significantly associated with worse progression-free survival (PFS both in univariate and multivariate analysis but is not associated with overall survival (n = 43 GBMs. PFS is a marker for responsiveness to treatment. We propose that TRADD-mediated NF-κB activation confers chemoresistance and thus a worse PFS in GBM. Consistent with the effect on PFS, silencing TRADD in glioma cells results in decreased NF-κB activity, decreased proliferation of cells, and increased sensitivity to temozolomide. TRADD expression is common in glioma-initiating cells. Importantly, silencing TRADD in GBM-initiating stem cell cultures results in decreased viability of stem cells, suggesting that TRADD may be required for maintenance of GBM stem cell populations. Thus, our study suggests that increased expression of cytoplasmic TRADD is both an important biomarker and a key driver of NF-κB activation in GBM and supports an oncogenic role for TRADD in GBM.

  6. Symmetrical infantile thalamic degeneration with focal cytoplasmic calcification.

    Science.gov (United States)

    Ambler, M; O'Neil, W

    1975-10-27

    Infantile thalamic degeneration is a rare clinico-pathological entity. Restricted location of the lesion and peculiar cytopathological changes serve to distinguish this disorder from other common encephalopathies. Optical and ultrastructural studies demonstrate cytoplasmic calcopherules in previously viable cells. According to current concepts of acute cellular reactions to injury and mechanism of intracellular calcification, the cytological changes cannot be attributed to either hypoxic ischemic cell change or dystrophic calcification. By analogy to other human and pathological material, the most likely basis for nondystrophic calcopherule formation is toxic or infectious injury with local synthesis, or autophagic or phagolysosomal degradation of cellular debris of specific chemical composition favoring calcium deposition.

  7. Drug-Induced Bullous Sweet Syndrome with Multiple Autoimmune Features

    Directory of Open Access Journals (Sweden)

    Jared J. Lund

    2010-01-01

    Full Text Available Sweet syndrome (SS (Acute Febrile Neutrophilic Dermatosis has been reported in association with autoimmune phenomena including relapsing polychondritis, drug-induced lupus, and the development of antineutrophil cytoplasmic antibodies (ANCAs. However, a combination of these autoimmune features has not been reported. Herein, we report a case of drug-induced bullous SS with ocular and mucosal involvement, glomerulonephritis, and multiple autoimmune features including clinical polychondritis with antitype II collagen antibodies, ANCAs, antinuclear (HEp-2, and antihistone antibodies in a patient on hydralazine and carbamazepine.

  8. An elderly ‘kawara’ craftsman with acute kidney injury and haemoptysis: a case of silica-induced autoimmunity

    OpenAIRE

    Matsui, Satoshi; Tsuji, Hiroko; Ono, Shinji

    2010-01-01

    A 62-year-old Japanese ‘kawara’ (ceramic roof tile) craftsman presented with acute kidney injury and haemoptysis. This case met the systemic lupus erythematosus and microscopic polyangiitis criteria, with high titres of myeloperoxidase–antineutrophil cytoplasmic antibody (570 EU). Results showed the presence of antinuclear antibody at a high titre (1:2560), but detection of rheumatoid factor, anti-dsDNA, anti-SSA and anti-SSB antibodies was not apparent. This serology was similar to drug-indu...

  9. Stage-specific appearance of cytoplasmic microtubules around the surviving nuclei during the third prezygotic division of Paramecium

    Institute of Scientific and Technical Information of China (English)

    Yiwen WANG; Jinqiang YUAN; Xin GAO; Xianyu YANG

    2012-01-01

    There are six micronuclear divisions during conjugation of Paramecium caudatum: three prezygotic and three postzygotic divisions.Four haploid nuclei are formed during the first two meiotic prezygotic divisions.Usually only one meiotic product is located in the paroral cone (PC) region at the completion of meiosis,which survives and divides mitotically to complete the third prezygotic division to yield a stationary and a migratory pronucleus.The remaining three located outside of the PC degenerate.The migratory pronuclei are then exchanged between two conjugants and fuse with the stationary pronuclei to form synkarya,which undergo three successive divisions (postzygotic divisions).However,little is known about the surviving mechanism of the PC nuclei.In the current study,stage-specific appearance of cytoplasmic microtubules (cMTs) was indicated during the third prezygotic division by immunofluorescence labeling with anti-alpha tubulin antibodies surrounding the surviving nuclei,including the PC nuclei and the two types of prospective pronuclei.This suggested that cMTs were involved in the formation of a physical barrier,whose function may relate to sequestering and protecting the surviving nuclei from the major cytoplasm,where degeneration of extra-meiotic products occurs,another important nuclear event during the third prezygotic division.

  10. Dynamic distribution of Ser-10 phosphorylated histone H3 in cytoplasm of MCF-7 and CHO cells during mitosis

    Institute of Scientific and Technical Information of China (English)

    Deng Wen LI; Qin YANG; Jia Tong CHEN; Hao ZHOU; Ru Ming LIU; Xi Tai HUANG

    2005-01-01

    The dynamic distribution of phosphorylated Histone H3 on Ser10 (phospho-H3) in cells was investigated to determine its function during mitosis. Human breast adenocarcinoma cells MCF-7, and Chinese hamster cells CHO were analyzed by indirect immunofluorescence staining with an antibody against phospho-H3. We found that the phosphorylation begins at early prophase, and spreads throughout the chromosomes at late prophase. At metaphase, most of the phosphoH3 aggregates at the end of the condensed entity of chromosomes at equatorial plate. During anaphase and telophase,the fluorescent signal of phospho-H3 is detached from chromosomes into cytoplasm. At early anaphase, phospho-H3shows ladder bands between two sets of separated chromosome, and forms "sandwich-like structure" when the chromosomes condensed. With the cleavage progressing, the "ladders" of the histone contract into a bigger bright dot. Then the histone aggregates and some of compacted microtubules in the midbody region are composed into a "bar-like"complex to separate daughter cells. The daughter cells seal their plasma membrane along with the ends of the "bar",inside which locates microtubules and modified histones, to finish the cytokinesis and keep the "bar complex" out of the cells. The specific distribution and kinetics of phospho-H3 in cytoplasm suggest that the modified histones may take part in the formation of midbody and play a crucial role in cytokinesis.

  11. Cytoplasmic and membranous CD24 marker expression has indirect correlation with cAMP/cGMP ratio

    Directory of Open Access Journals (Sweden)

    Hossein Goudarzi

    2014-02-01

    Full Text Available Background- CD24 is a cell adhesion molecule that has been implicated in metastatic tumor progression cells. Our aim was clarify correlation between CD24 expression and cAMP:cGMP ratio in murine colorectal cancer cell line (CT26 after using cholera toxin.Materials and Methods- The CT26 cells were cultured in microtubes for assaying cAMP and cGMP; also the cells were cultured in flasks for assaying cytoplasmic and membranous CD24 expression. The Real-Time PCR was done for cDNA that was synthesized from CT26 cells’ mRNA. Also, expression CD24 marker of cells was determined by Anti-CD24 antibody and Goat Anti-Rabbit IgG-FITC (flow cytometry.Results- The cholera toxin grew cAMP:cGMP ratio and it influenced cytoplasmic and membrane CD24 expression.Conclusion- There are indirect correlation between cAMP:cGMP ratio and CD24 expression.

  12. Effects of cytoplasmic inheritance on preweaning traits of Hereford cattle

    Directory of Open Access Journals (Sweden)

    Mezzadra Carlos Alberto

    2005-01-01

    Full Text Available The influence of cytoplasmic inheritance on birth and weaning weight was evaluated in an experimental Hereford herd. Data on 1,720 records for birth and weaning weights from calves born between 1963 and 2002 were studied. Variance components were estimated using MTDFREML procedures and an animal model was fitted for each trait. Direct and maternal additive effects and permanent environment and maternal lineage effects were treated as random, while year and month of birth, age of dam and sex of the calf were treated as fixed. Identification of maternal lineages was based on pedigree information. The contribution to phenotypic variance of cytoplasmic lineages defined by pedigree information was negligible for both traits. Mitochondrial genotypes of cows present in the herd in 2002 were analyzed by single strand conformation polymorphism (SSCP analysis. Only five different genotypes were identified among 23 maternal lineages. All the animals with records were assigned to maternal genotypes based on pedigree information. The statistical analysis was repeated, removing maternal lineage from the model and including mitochondrial genotype as a fixed effect. No evidence of genotype effects was detected. These results suggest a negligible effect of the mitochondrial genome on the preweaning traits of this Hereford herd.

  13. CryoEM structure of yeast cytoplasmic exosome complex.

    Science.gov (United States)

    Liu, Jun-Jie; Niu, Chu-Ya; Wu, Yao; Tan, Dan; Wang, Yang; Ye, Ming-Da; Liu, Yang; Zhao, Wenwei; Zhou, Ke; Liu, Quan-Sheng; Dai, Junbiao; Yang, Xuerui; Dong, Meng-Qiu; Huang, Niu; Wang, Hong-Wei

    2016-07-01

    The eukaryotic multi-subunit RNA exosome complex plays crucial roles in 3'-to-5' RNA processing and decay. Rrp6 and Ski7 are the major cofactors for the nuclear and cytoplasmic exosomes, respectively. In the cytoplasm, Ski7 helps the exosome to target mRNAs for degradation and turnover via a through-core pathway. However, the interaction between Ski7 and the exosome complex has remained unclear. The transaction of RNA substrates within the exosome is also elusive. In this work, we used single-particle cryo-electron microscopy to solve the structures of the Ski7-exosome complex in RNA-free and RNA-bound forms at resolutions of 4.2 Å and 5.8 Å, respectively. These structures reveal that the N-terminal domain of Ski7 adopts a structural arrangement and interacts with the exosome in a similar fashion to the C-terminal domain of nuclear Rrp6. Further structural analysis of exosomes with RNA substrates harboring 3' overhangs of different length suggests a switch mechanism of RNA-induced exosome activation in the through-core pathway of RNA processing. PMID:27174052

  14. Neutralization resistance of virological synapse-mediated HIV-1 Infection is regulated by the gp41 cytoplasmic tail.

    Science.gov (United States)

    Durham, Natasha D; Yewdall, Alice W; Chen, Ping; Lee, Rebecca; Zony, Chati; Robinson, James E; Chen, Benjamin K

    2012-07-01

    Human immunodeficiency virus type 1 (HIV-1) infection can spread efficiently from infected to uninfected T cells through adhesive contacts called virological synapses (VSs). In this process, cell-surface envelope glycoprotein (Env) initiates adhesion and viral transfer into an uninfected recipient cell. Previous studies have found some HIV-1-neutralizing patient sera to be less effective at blocking VS-mediated infection than infection with cell-free virus. Here we employ sensitive flow cytometry-based infection assays to measure the inhibitory potency of HIV-1-neutralizing monoclonal antibodies (MAb) and HIV-1-neutralizing patient sera against cell-free and VS-mediated infection. To various degrees, anti-Env MAbs exhibited significantly higher 50% inhibitory concentration (IC(50)s) against VS-mediated infection than cell-free infection. Notably, the MAb 17b, which binds a CD4-induced (CD4i) epitope on gp120, displayed a 72-fold reduced efficacy against VS-mediated inocula compared to cell-free inocula. A mutant with truncation mutation in the gp41 cytoplasmic tail (CT) which is unable to modulate Env fusogenicity in response to virus particle maturation but which can still engage in cell-to-cell infection was tested for the ability to resist neutralizing antibodies. The ΔCT mutation increased cell surface staining by neutralizing antibodies, significantly enhanced neutralization of VS-mediated infection, and had reduced or no effect on cell-free infection, depending upon the antibody. Our results suggest that the gp41 CT regulates the exposure of key neutralizing epitopes during cell-to-cell infection and plays an important role in immune evasion. Vaccine strategies should consider immunogens that reflect Env conformations exposed on the infected cell surface to enhance protection against VS-mediated HIV-1 spread. PMID:22553332

  15. HIV Antibody Test

    Science.gov (United States)

    ... be limited. Home Visit Global Sites Search Help? HIV Antibody and HIV Antigen (p24) Share this page: Was this page helpful? Also known as: HIV Screening Tests; AIDS Test; AIDS Screen; HIV Serology; ...

  16. Antinuclear antibody panel

    Science.gov (United States)

    ... blood may be due to: Chronic liver disease Collagen vascular disease Drug-induced lupus erythematosus Myositis (inflammatory muscle disease) ... Saunders; 2011:chap 51. Read More Antibody Arthritis Collagen vascular disease Drug-induced lupus erythematosus Liver disease Scleroderma Systemic ...

  17. Anti-cartilage antibody.

    Science.gov (United States)

    Greenbury, C L; Skingle, J

    1979-08-01

    Antibody to cartilage has been demonstrated by indirect immunofluorescence on rat trachea in the serum of about 3% of 1126 patients with rheumatoid arthritis. Titres ranged from 1:20 to 1:640. The antibody was not found in 284 patients with primary or secondary osteoarthritis or in 1825 blood donors, nor, with the exception of two weak reactors, in 1314 paraplegic patients. In most cases the antibody appears to be specific for native type II collagen. Using this as an antigen in a haemagglutination test 94% of anti-cartilage sera were positive, whereas among 100 rheumatoid control sera there were only three weak positives. More than 80% of patients with antibody had some erosion of articular cartilage, but there was no correlation with age, sex, duration of disease, nor any recognisable clinical event or change.

  18. Antibody tumor penetration

    Science.gov (United States)

    Thurber, Greg M.; Schmidt, Michael M.; Wittrup, K. Dane

    2009-01-01

    Antibodies have proven to be effective agents in cancer imaging and therapy. One of the major challenges still facing the field is the heterogeneous distribution of these agents in tumors when administered systemically. Large regions of untargeted cells can therefore escape therapy and potentially select for more resistant cells. We present here a summary of theoretical and experimental approaches to analyze and improve antibody penetration in tumor tissue. PMID:18541331

  19. Expression of Recombinant Antibodies

    OpenAIRE

    Frenzel, André; Hust, Michael; Schirrmann, Thomas

    2013-01-01

    Recombinant antibodies are highly specific detection probes in research, diagnostics, and have emerged over the last two decades as the fastest growing class of therapeutic proteins. Antibody generation has been dramatically accelerated by in vitro selection systems, particularly phage display. An increasing variety of recombinant production systems have been developed, ranging from Gram-negative and positive bacteria, yeasts and filamentous fungi, insect cell lines, mammalian cells to transg...

  20. Skeletal Phenotype of Transgenic Mice Expressing the Beta1 Integrin Cytoplasmic Tail In Osteoblasts

    Science.gov (United States)

    Globus, R. K.; vanderMeulen, M. C. H.; Damsky, D.; Kim, J.-B.; Amblard, D.; Amblard, D.; Nishimura, Y.; Almeida, E.; Iwaniec, U. T.; Wronski, T. J.; Dalton, Bonnie (Technical Monitor)

    2002-01-01

    To define the physiologic role of beta1 integrin in bone formation and mechanical loading, transgenic mice were generated by expressing the cytoplasmic tall and transmembrane domain of Beta1 integrin under the control of the osteocalcin promoter. In cultured cells, this truncated fragment of Beta1 can act as a dominant negative. Previously, the matrix of calvariae was shown to be abnormal in transgenic (TG) compared to wildtype (WT) mice. In this study, we analyzed appendicular bone in TG and WT, male and female mice at 14, 35, 63, 90 and 365 days old (n=8-12/gp). To assess beta1 integrin function in mechanical loading, a pilot study using hindlimb unloading by tail suspension was performed. 35d old TG and WT females were hindlimb unloaded for 4 wks (n=3-5). Body mass, bone mineral content, histomorphometric (distal femur) and biomechanical parameters were analyzed. Statistical significance (P less than.05) was defined by ANOVA using the Tukey-Kramer post-hoc test. We confirmed transgene expression by immunoprecipitating then immunoblotting bone lysates using an antibody against the beta1 tail. Body masses of TG mice at 63, 90 and 365d old were greater (16-25%) than WT. Some TG female mice at 365d appeared obese; mean abdominal fat mass was 415% greater in TG than WT mice. Tibiae were longer (5-7%) in TG than WT mice at 63 and 90d. Tibial mineral mass of 35d males was 7% lower in TG than WT mice, but at 63d was 21% higher. The % osteoblast surface in 35d TG mice was 20% higher than WT, and at 63d was 17% lower, while % osteoclast surface did not differ. In 365d mice, cancellous bone volume (125%) and endocortical mineral apposition rate (40%) were greater in TG than WT males but not females. In WT mice, hindlimb unloading caused a reduction in mineral mass of tibiae (-20%) and lumbar vertebrae (-22%) relative to normally loaded controls. Surprisingly, hindlimb unloading also caused a relative reduction (-13%) in humerus mass. The effects of hindlimb unloading on

  1. Antibody informatics for drug discovery

    DEFF Research Database (Denmark)

    Shirai, Hiroki; Prades, Catherine; Vita, Randi;

    2014-01-01

    for antibody rational design using computational approaches to affinity and stability improvement, as well as ab-initio and homology-based antibody modeling; (ii) resources for antibody sequences, structures, and immune epitopes and open drug discovery resources for development of antibody drugs; and (iii...

  2. Actin and myosin regulate cytoplasm stiffness in plant cells: a study using optical tweezers.

    Science.gov (United States)

    van der Honing, Hannie S; de Ruijter, Norbert C A; Emons, Anne Mie C; Ketelaar, Tijs

    2010-01-01

    Here, we produced cytoplasmic protrusions with optical tweezers in mature BY-2 suspension cultured cells to study the parameters involved in the movement of actin filaments during changes in cytoplasmic organization and to determine whether stiffness is an actin-related property of plant cytoplasm. Optical tweezers were used to create cytoplasmic protrusions resembling cytoplasmic strands. Simultaneously, the behavior of the actin cytoskeleton was imaged. After actin filament depolymerization, less force was needed to create cytoplasmic protrusions. During treatment with the myosin ATPase inhibitor 2,3-butanedione monoxime, more trapping force was needed to create and maintain cytoplasmic protrusions. Thus, the presence of actin filaments and, even more so, the deactivation of a 2,3-butanedione monoxime-sensitive factor, probably myosin, stiffens the cytoplasm. During 2,3-butanedione monoxime treatment, none of the tweezer-formed protrusions contained filamentous actin, showing that a 2,3-butanedione monoxime-sensitive factor, probably myosin, is responsible for the movement of actin filaments, and implying that myosin serves as a static cross-linker of actin filaments when its motor function is inhibited. The presence of actin filaments does not delay the collapse of cytoplasmic protrusions after tweezer release. Myosin-based reorganization of the existing actin cytoskeleton could be the basis for new cytoplasmic strand formation, and thus the production of an organized cytoarchitecture.

  3. Engineering antibodies for cancer therapy.

    Science.gov (United States)

    Boder, Eric T; Jiang, Wei

    2011-01-01

    The advent of modern antibody engineering has led to numerous successes in the application of these proteins for cancer therapy in the 13 years since the first Food and Drug Administration approval, which has stimulated active interest in developing more and better drugs based on these molecules. A wide range of tools for discovering and engineering antibodies has been brought to bear on this challenge in the past two decades. Here, we summarize mechanisms of monoclonal antibody therapeutic activity, challenges to effective antibody-based treatment, existing technologies for antibody engineering, and current concepts for engineering new antibody formats and antibody alternatives as next generation biopharmaceuticals for cancer treatment.

  4. Cytoplasmic sulfur trafficking in sulfur-oxidizing prokaryotes.

    Science.gov (United States)

    Dahl, Christiane

    2015-04-01

    Persulfide groups are chemically versatile and participate in a wide array of biochemical pathways. Although it is well documented that persulfurated proteins supply a number of important and elaborate biosynthetic pathways with sulfane sulfur, it is far less acknowledged that the enzymatic generation of persulfidic sulfur, the successive transfer of sulfur as a persulfide between multiple proteins, and the oxidation of sulfane sulfur in protein-bound form are also essential steps during dissimilatory sulfur oxidation in bacteria and archaea. Here, the currently available information on sulfur trafficking in sulfur oxidizing prokaryotes is reviewed, and the idea is discussed that sulfur is always presented to cytoplasmic oxidizing enzymes in a protein-bound form, thus preventing the occurrence of free sulfide inside of the prokaryotic cell. Thus, sulfur trafficking emerges as a central element in sulfur-oxidizing pathways, and TusA homologous proteins appear to be central and common elements in these processes.

  5. Cytoplasmic flows as signatures for the mechanics of mitotic positioning

    CERN Document Server

    Nazockdast, Ehssan; Needleman, Daniel; Shelley, Michael

    2015-01-01

    The proper positioning of the mitotic spindle is crucial for asymmetric cell division and generating cell diversity during development. Proper position in the single-cell embryo of Caenorhabditis elegans is achieved initially by the migration and rotation of the pronuclear complex (PNC) and its two associated centrosomal arrays of microtubules (MTs). We present here the first systematic theoretical study of how these $O(1000)$ centrosomal microtubules (MTs) interact through the immersing cytoplasm, the cell periphery and PNC, and with each other, to achieve proper position. This study is made possible through our development of a highly efficient and parallelized computational framework that accounts explicitly for long-ranged hydrodynamic interactions (HIs) between the MTs, while also capturing their flexibility, dynamic instability, and interactions with molecular motors and boundaries. First, we show through direct simulation that previous estimates of the PNC drag coefficient, based on either ignoring or ...

  6. Cytoplasm T-maize tissue culture to obtain methomyl resistance

    International Nuclear Information System (INIS)

    The calli of two cytoplasm T-maize hybrids were cultured in vitro with methomyl in the medium in order to obtain resistance. A mutagenic pretreatment was performed in one of the hybrids by injecting ethylmethane sulphonate (EMS) into the ear cob before fetilization. The atrophic tissue method was utilized to obtain totipotent callus. After treatment, coloured pericarp grains were observed in the regenerated plants. Different esterase electrophoretic patterns also appeared in one of them. In the methomyl resistance test, some nearly resistant (R-) and some less susceptible (S-) plants were detected within the same descendance (possibly due to mitochondrial variability). The interaction between mutagenic treatment with EMS and in vitro culture may be a suitable method for inducing changes of methomyl susceptibility in cms-T maize plants. (author)

  7. Genetics of Fertility Restoration in Cytoplasmic Male Sterile Pepper

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Pepper hybrid seeds production using male sterility could lower cost by reducing time and labour, and increase the genetic purity of the F1 seeds. To investigate the genetics of fertility restoration of the Peterson cytoplasmic sterility in pepper, a doubled haploid population of 115 pepper lines obtained from anther culture of the F1 hybrid between Yolo Wonder (sterility maintainer line) and Perennial (fertility restorer line) and the parental lines were test-crossed by 77013A (a strict cytoplasmic-genic male sterile line). The fertility of the test-crossed lines was assessed in greenhouse and open field with the following three criteria: pollen index (PI, visual estimation of pollen amount per flower), pollen number (PN, pollen counting under microscope), and seed number (SN, the number of seeds per fruit in open pollination). Correlations between the each couple of criteria within, as well as between the cultivation methods ranged from 0.55 to 0.84. Analysis of variance showed that the genotype (DH line) and environment were the significant sources of variation of the fertility.Narrow sense of heritance of fertility restoration ranged from 0.38 to 0.92, depending on the criteria and environment. The distribution of the progeny was continuous between the parental genotypes indicating the quantitative inheritance of fertility restoration. Inferred from segregation according to Snape et al.(1984), the number of segregating genes was estimated to be that three to four genetic factors were involved in pollen traits (PI and PN) and five to eight genetic factors in seed production (SN). The heredity analysis of the CMS will be helpful for understanding of the genetic mechanism of the fertility restoration and the exploitation of the CMS in hybrid seed production.

  8. Regulation of Cytoplasmic Dynein ATPase by Lis1

    Science.gov (United States)

    Mesngon, Mariano T.; Tarricone, Cataldo; Hebbar, Sachin; Guillotte, Aimee M.; Schmitt, E. William; Lanier, Lorene; Musacchio, Andrea; King, Stephen J.; Smith, Deanna S.

    2015-01-01

    Mutations in Lis1 cause classical lissencephaly, a developmental brain abnormality characterized by defects in neuronal positioning. Over the last decade, a clear link has been forged between Lis1 and the microtubule motor cytoplasmic dynein. Substantial evidence indicates that Lis1 functions in a highly conserved pathway with dynein to regulate neuronal migration and other motile events. Yeast two-hybrid studies predict that Lis1 binds directly to dynein heavy chains (Sasaki et al., 2000; Tai et al., 2002), but the mechanistic significance of this interaction is not well understood. We now report that recombinant Lis1 binds to native brain dynein and significantly increases the microtubule-stimulated enzymatic activity of dynein in vitro. Lis1 does this without increasing the proportion of dynein that binds to microtubules, indicating that Lis1 influences enzymatic activity rather than microtubule association. Dynein stimulation in vitro is not a generic feature of microtubule-associated proteins, because tau did not stimulate dynein. To our knowledge, this is the first indication that Lis1 or any other factor directly modulates the enzymatic activity of cytoplasmic dynein. Lis1 must be able to homodimerize to stimulate dynein, because a C-terminal fragment (containing the dynein interaction site but missing the self-association domain) was unable to stimulate dynein. Binding and colocalization studies indicate that Lis1 does not interact with all dynein complexes found in the brain. We propose a model in which Lis1 stimulates the activity of a subset of motors, which could be particularly important during neuronal migration and long-distance axonal transport. PMID:16481446

  9. [Pulmonar pseudotumor in granulomatosis with polyangiitis (GPA). Pulmonary cancer and/or GPA? Diagnostic implications of pulmonary nodules].

    Science.gov (United States)

    Horta-Baas, Gabriel; Meza-Zempoaltecatl, Esteban; Pérez-Cristóbal, Mario; Barile-Fabris, Leonor Adriana

    2016-01-01

    Granulomatosis with polyangiitis (GPA), formerly known as Wegener's granulomatosis, is a systemic necrotizing vasculitis, which affects small and medium sized blood vessels and is often associated with cytoplasmic anti-neutrophil cytoplasmic antibodies (ANCA). Inflammatory pseudotumor is a rare condition characterized by the appearance of a mass lesion that mimics a malignant tumor both clinically and on imaging studies, but that is thought to have an inflammatory/reactive pathogenesis. We report a patient with a GPA which was originally diagnosed as malignancy. PMID:27595257

  10. MSL2 Promotes Mdm2-independent Cytoplasmic Localization of p53*

    OpenAIRE

    Kruse, Jan-Philipp; Gu, Wei

    2009-01-01

    Although it was originally thought of as a passive way to block the nuclear function of p53, accumulating evidence suggests that cytoplasmic localization of p53 plays an active role in p53-mediated functions such as apoptosis and autophagy. Previous studies by us and others demonstrated that Mdm2-mediated p53 ubiquitination induces both degradation and cytoplasmic localization. Here we describe MSL2, a novel E3 ligase for p53 that promotes ubiquitin-dependent cytoplasm...

  11. Studies on the Utilization Potentiality of the Nucleo-Cytoplasmic Hybrid in Wheat

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    A series of comparative studies was carried out on the genetic effects of 25 alien cytoplasms of wheat on the growth potential, heading stage , fertility, resistance against diseases, important agronomic traits and its heterosis of 125 nuclei-cytoplasmic hybrids of wheat. The results indicated that there were clearly effects of alien cytoplasms on some characteristics, but the nucleus still exerted main effect on other characteristics. The effect of interactions between nucleus and cytoplasm was comparative obvious in some combination. Consequently, when we utilize the effects of alien cytoplasms, we should pay full attention to the facts such as the characteristic to be improved, the effects of cytoplasm ,nucleus, the nucleus-cytoplasm interactions on that characteristics . From the preliminary studies, we believed that the cytoplasmic types of M°, S1, Sv, D2, D and B, and the nucleo-cytoplasmic hybrids of (Ae. sharonensis) -Bl74, (Ae. squarrosa)352-35, (Ae. cylindrica) -352-35, (Ae. cylindrica)-E EN-1, (Ae. cylindrica)- NPFP, and (Ae. speltoides)352-35 would have some utilization potentiality in cultivar improvement.

  12. Pollen mitochondria in cytoplasmically male sterile tobacco zygotic and embryonic cells

    International Nuclear Information System (INIS)

    An attempt is being made to establish cytoplasmic organelles transmission during the process of fertilization, by using tobacco grain pollen labelled with leucine 14C and tritiated thymidine. Through autoradiography the fate of pollen germination and its entry into the embryo sac has been studied. A few days after fertilization, labelled cytoplasmic organelles - mainly mitochondria - were detected in the embryo sac. However, labelling was not observed in cytoplasmic organelles by using tritiated thymidine. For more conclusive results labelled DNA incorporated in cytoplasmic organelles have to be traced during the embryo and endosperm development

  13. Antibody affinity maturation

    DEFF Research Database (Denmark)

    Skjødt, Mette Louise

    linker for yeast surface display of scFv and scFab fragments, we compared a series of different Gly-Ser-based linkers in display and antigen binding proficiency. We show that these formats of the model antibody can accommodate linkers of different lengths and that introduction of alanine or glutamate......-2. Based on the presented data we suggest that affinity maturation of the model antibody proceeds through multiple incremental steps of subtle improvements. We moreover conclude that the best affinity improved candidates are likely to be obtained through optimization of both the antigen...... fragments by in vivo homologous recombination large combinatorial antibody libraries can easily be generated. We have optimized ordered assembly of three CDR fragments into a gapped vector and observed increased transformation efficiency in a yeast strain carrying a deletion of the SGS1 helicase...

  14. Cytoplasmic p21WAF1/CIP1 expression is correlated with HER-2/ neu in breast cancer and is an independent predictor of prognosis

    International Nuclear Information System (INIS)

    HER-2 (c-erbB2/Neu) predicts the prognosis of and may influence treatment responses in breast cancer. HER-2 activity induces the cytoplasmic location of p21WAFI/CIPI in cell culture, accompanied by resistance to apoptosis. p21WAFI/CIPI is a cyclin-dependent kinase inhibitor activated by p53 to produce cell cycle arrest in association with nuclear localisation of p21WAFI/CIPI. We previously showed that higher levels of cytoplasmic p21WAFI/CIPI in breast cancers predicted reduced survival at 5 years. The present study examined HER-2 and p21WAFI/CIPI expression in a series of breast cancers with up to 9 years of follow-up, to evaluate whether in vitro findings were related to clinical data and the effect on outcome. The CB11 anti-HER2 monoclonal antibody and the DAKO Envision Plus system were used to evaluate HER-2 expression in 73 patients. p21WAFI/CIPI staining was performed as described previously using the mouse monoclonal antibody Ab-1 (Calbiochem, Cambridge, MA, USA). HER-2 was evaluable in 67 patients and was expressed in 19% of cases, predicting reduced overall survival (P = 0.02) and reduced relapse-free survival (P = 0.004; Cox regression model). HER-2-positive tumours showed proportionately higher cytoplasmic p21WAFI/CIPI staining using an intensity distribution score (median, 95) compared with HER-2-negative cancers (median, 47) (P = 0.005). There was a much weaker association between nuclear p21WAFI/CIPI and HER-2 expression (P = 0.05), suggesting an inverse relationship between nuclear p21WAF1/CIP1 and HER-2. This study highlights a new pathway by which HER-2 may modify cancer behaviour. HER-2 as a predictor of poor prognosis may partly relate to its ability to influence the relocalisation of p21WAFI/CIPI from the nucleus to the cytoplasm, resulting in a loss of p21WAFI/CIPItumour suppressor functions. Cytoplasmic p21WAFI/CIPI may be a surrogate marker of functional HER-2 in vivo

  15. Expression of anti-SRP19 antibody in muscle tissues from patients with autoimmune necrotizing myopathy.

    Science.gov (United States)

    Wang, Q; Duan, F; Liu, P; Wang, P F; Wang, M X

    2016-01-01

    This study aimed to investigate the role of anti-SRP19 antibody in muscle tissues of patients with autoimmune necrotizing myopathy. Immunohistochemistry staining was used to determine the expression of anti-SRP19 antibodies in muscle tissues of autoimmune necrotizing myopathy patients. Results demonstrated that anti-SRP19 antibody was expressed in 71.4% (20/28) of muscle tissue specimens from patients with autoimmune necrotizing myopathy. Anti-SRP19 antibody expression was mainly localized in cytoplasm of necrotic muscle fibers surrounding the small blood vessels and interstitial cells. There were no significant differences in the age, course of disease, muscle, and creatine kinase levels between patients with positive or negative expression of anti-SRP19 antibodies. The expression levels of anti-SRP19, serum anti-nuclear antibodies, as well as anti-Ro-52, anti- SSA, anti-Sm, and anti-Jo-1 antibodies were not significantly different among groups. This study demonstrates that anti-SRP19 antibody is highly expressed in muscle tissues of patients with autoimmune necrotizing myopathy, and suggests that this protein may be involved in the origin and progression of the disease. PMID:27525944

  16. Antithyroglobulin Antibodies and Antimicrosomal Antibodies in Various Thyroid Diseases

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Gwon Jun; Hong, Key Sak; Choi, Kang Won; Lee, Kyu; Koh, Chang Soon; Lee, Mun Ho; Park, Sung Hoe; Chi, Je Geun; Lee, Sang Kook [Seoul National University College of Medicine, Seoul (Korea, Republic of)

    1979-03-15

    The authors investigated the incidence of antithyroglobulin antibodies and antibodies and antimicrosomal antibodies measured by tanned red cell hemagglutination method in subjects suffering from various thyroid disorders. 1) In 15 normal patients, neither suffering from any thyroid diseases nor from any other autoimmune disorders, the antithyroglobulin antibodies were all negative, but the antimicrosomal antibody was positive only in one patient (6.7%). 2) The antithyroglobulin antibodies were positive in 31.5% (34 patients) of 108 patients with various thyroid diseases, and the antimicrosomal antibodies were positive in 37.0% (40 patients). 3) of the 25 patients with Graves' diseases, 7 patients (28.0%) showed positive for the antithyroglobulin antibodies, and 9 (36.0%) for the antimicrosomal antibodies. There was no definite differences in clinical and thyroid functions between the groups with positive and negative results. 4) Both antibodies were positive in 16 (88.9%) and 17 (94.4%) patients respectively among 18 patients with Hashimoto's thyroiditis, all of them were diagnosed histologically. 5) Three out of 33 patients with thyroid adenoma showed positive antibodies, and 3 of 16 patients with thyroid carcinoma revealed positive antibodies. 6) TRCH antibodies demonstrated negative results in 2 patients with subacute thyroiditis, but positive in one patient with idiopathic primary myxedema. 7) The number of patients with high titers(>l:802) was 16 for antithyroglobulin antibody, and 62.5% (10 patients) of which was Hashimoto's thyroiditis. Thirteen (65.0) of 20 patients with high titers (>l:802) for antimicrosomal antibody was Hashimoto's thyroiditis. TRCH test is a simple, sensitive method, and has high reliability and reproducibility. The incidences and titers of antithyroglobulin antibody and antimicrosomal antibody are especially high in Hashimoto's thyroiditis.

  17. Distinct Cytoplasmic Expression of KL-6 Mucin in Chromophobe Renal Cell Carcinoma: A Comparative Immunohistochemical Study with Other Renal Epithelial Cell Tumors

    International Nuclear Information System (INIS)

    The presence of cytoplasmic sialyl glycoproteins is a conspicuous feature in chromophobe renal cell carcinoma (RCC). We compared the immunohistochemical expression of sialyl glycoproteins in chromophobe RCC with that in other types of renal tumors. Formalin-fixed, paraffin-embedded tissues of surgically resected renal tumors (chromophobe RCC, 14 cases [10 cases of classic type and 4 cases of eosinophilic variant]; oncocytoma, 7 cases; and clear cell RCC, 9 cases) and kidneys from immature infants (4 cases) were immunostained with antibodies against sialyl glycoproteins (anti-KL-6 and anti-sialyl MUC1 antibodies). Cytoplasmic expression of KL-6 and sialyl MUC1 was distinctive in the chromophobe RCC and renal oncocytoma cells, and in the intercalated cells in collecting duct epithelia. Apical-surface staining of these sialyl glycoproteins was predominantly observed in clear RCC, in the epithelia of the distal tubule and collecting duct, and in the neonatal renal proximal tubule, but not in those of the adult renal proximal tubule. The above-mentioned observations provide additional evidence for similar phenotypic profiles of chromophobe RCC and renal oncocytoma, and the intercalated cells in collecting ducts and the oncofetal expression of sialyl glycoproteins in clear cell RCC. KL-6 is a potential tumor marker for renal tumors

  18. Effects of alien and intraspecies cytoplasms on manifestation of nuclear genes for wheat resistance to brown rust: II. Specificity of cytoplasm influence on different Lr genes

    Energy Technology Data Exchange (ETDEWEB)

    Voluevich, E.A.; Buloichik, A.A.; Palilova, A.N. [Institute of Genetics and Cytology, Minsk (Belarus)

    1995-04-01

    Specificity of expression of the major nuclear genes Lr to two brown rust clones in hybrids with the same maternal cytoplasm was analyzed. It was evaluated by a resistant: susceptible ratio in the F{sub 2}. Reciprocal hybrids were obtained from the cross between the progeny of homozygous susceptible plants of the cultivar Penjamo 62 and its alloplasmatic lines carrying cytoplasms of Triticum dicoccoides var. fulvovillosum, Aegilops squarrosa var. typical, Agropyron trichophorum, and isogenic lines of the cultivar Thatcher (Th) with the Lr1, Lr9, Lr15, and Lr19 genes. It was shown that the effect of the Lr1 gene in the cytoplasm of cultivar Thatcher and in eu-, and alloplasmatic forms of Penjamo 62 was less expressed than that of other Lr genes. Cytoplasm of the alloplasmatic line (dicoccoides)-Penjamo 62 was the only exception: in the F{sub 2}, hybrids with Th (Lr1) had a higher yield of resistant forms than those with Th (Lr15). In the hybrid combinations studied, expression and/or transmission of the Lr19 gene was more significant than that of other genes. This gene had no advantages over Lr15 and Lr19 only in cytoplasm of the alloplasmatic line (squarrosa)-Penjamo 62. In certain hybrid cytoplasms, the display of the Lr1, Lr15, and Lr19 genes, in contrast to Lr9, varied with the virulence of the pathogen clones. 15 refs., 5 tabs.

  19. Responses of alloplasmic (cytoplasm=Triticum timopheevii) and euplasmic wheats (Triticum aestivum) to photoperiod and vernalization.

    Science.gov (United States)

    Ward, R W; Heyne, E G; Paulsen, G M

    1983-07-01

    Studies were conducted to determine the influence of the male sterility-inducing cytoplasm of Triticum timopheevii (Zhuk.) Zhuk. on response of several common winter wheat (T. aestivum L.) nuclear genotypes to photoperiod and vernalization. Comparative studies of cytoplasmic substitution lines provide information on the role of the cytoplasmic genetic mechanism in growth and development. In the case of cytoplasmic male sterility-based hybrid production systems, ubiquity of sterility-inducing cytoplasm in derived hybrids warrants thorough characterization of its influence on plant phenotype. Factorial combinations of cytoplasm (T. timopheevii and T. aestivum), nuclear genotype, and photoperiod or vernalization treatments were evaluated under hydroponic conditions in controlled environment chambers. Interaction of cytoplasm, photoperiod, and nuclear genotype was significant in one or more experiments for days to anthesis and potential spikelet number, and interaction of cytoplasm, vernalization, and nuclear genotype was significant for days to spike emergence. Long day length was associated with increased percentage seed set in one study, but interactions of photoperiod and cytoplasm were not detected for percentage seed set. Interactions involving cytoplasm and photoperiod or vernalization were interpreted as evidence of the existence of genetic factors in cytoplsam of T. timopheevii which alter photoperiod or vernalization responses of alloplasmic plants relative to responses exhibited by euplasmic plants. Since photoperiod and vernalization responses are critical to adaptation, T. timopheevii cytoplasm can alter adaptability of T. aestivum. The specific effect would be nuclear genotype dependent, and does not appear to be of a magnitude greater than that induced by nuclear genetic variability at loci conditioning photoperiod or vernalization responses or other adaptation-determining characteristics. Normal multilocation/year testing of alloplasmic hybrids should

  20. Genetic diagnosis of cytoplasmic male sterile cybrid plants of rice.

    Science.gov (United States)

    Akagi, H; Nakamura, A; Sawada, R; Oka, M; Fujimura, T

    1995-06-01

    Twelve Japanese rice cultivars were converted to CMS by asymmetric protoplast fusion with MTC-5A, the cytoplasm of which was derived from an indica rice, Chinsurah Boro II. With the exception of the cybrids that had a nucleus from Hoshiyutaka, most of these cybrid plants were sterile. The unique sequence downstream from the mitochondrial atp6 of MTC-5A was specifically amplified in the sterile cybrid plants by PCR. All progenies of the cybrid plants carrying this unique sequence were sterile. On the other hand, in some of the sterile cybrid plants in which the unique sequence was not amplified by PCR, fertility was recovered in their progenies. Somaclonal mutation may have caused sterility in these cybrids. Only the cybrid plants that had the unique sequence detected by PCR were CMS. Thus, the CMS plants can be selected rapidly and easily by PCR, at an early stage of plant regeneration. Soon after transplanting the regenerated plants to a green house, fertile cybrids and sterile cybrids produced by somaclonal mutation can be removed. These findings also show that the unique region downstream from atp6 is tightly linked with the CMS phenotype. PMID:24173048

  1. Genetic and molecular basis of cytoplasmic male sterility in maize

    Directory of Open Access Journals (Sweden)

    Parvez A. Sofi

    2007-08-01

    Full Text Available Cytoplasmic male sterility is a maternally inherited trait that suppresses pollen production due to the interaction of nuclear and mitochondrial genomes. In maize three types of CMS systems, namely CMS-T, CMS-S and CMS-C, have been documented and are differentiated by the reaction to restorers, mitochondrial DNA restriction digest patterns, and complements of low molecular weight plasmids. CMS-T is restored fully by Rf-1 and Rf-2, CMS-S by Rf-3, and CMS-C by Rf-4. All restorer genes except Rf-2 restore fertility by affecting the transcript profile of CMS-associated locus. The sterility is caused by the disorganization of the tapetum and surrounding cell layers as a result of the expression of pollen specific genes. Even though such phenotypes are associated with gene dysfunction in mitochondria, the chloroplasts have emerged as ideal organs for engineering male sterility in crop plants. A number of systems such as barnase-barstar have been standardized in Brassica. Recently, polyhydroxy butyrate was identified as a potential candidate gene for engineering male sterility. Moreover, a broad group of proteins called PPR (pentatricopeptide repeat proteins has also shown to hold great promise for engineering male sterility in crop plants as most of the restorers belong to this category. In maize one such protein, CRP-1, has been identified

  2. In silico classification of proteins from acidic and neutral cytoplasms.

    Directory of Open Access Journals (Sweden)

    Yaping Fang

    Full Text Available Protein acidostability is a common problem in biopharmaceutical and other industries. However, it remains a great challenge to engineer proteins for enhanced acidostability because our knowledge of protein acidostabilization is still very limited. In this paper, we present a comparative study of proteins from bacteria with acidic (AP and neutral cytoplasms (NP using an integrated statistical and machine learning approach. We construct a set of 393 non-redundant AP-NP ortholog pairs and calculate a total of 889 sequence based features for these proteins. The pairwise alignments of these ortholog pairs are used to build a residue substitution propensity matrix between APs and NPs. We use Gini importance provided by the Random Forest algorithm to rank the relative importance of these features. A scoring function using the 10 most significant features is developed and optimized using a hill climbing algorithm. The accuracy of the score function is 86.01% in predicting AP-NP ortholog pairs and is 76.65% in predicting non-ortholog AP-NP pairs, suggesting that there are significant differences between APs and NPs which can be used to predict relative acidostability of proteins. The overall trends uncovered in the study can be used as general guidelines for designing acidostable proteins. To best of our knowledge, this work represents the first systematic comparative study of the acidostable proteins and their non-acidostable orthologs.

  3. Model for Unidirectional Movement of Axonemal and Cytoplasmic Dynein Molecules

    Institute of Scientific and Technical Information of China (English)

    Ping XIE; Shuo-Xing DOU; Peng-Ye WANG

    2006-01-01

    A model for the unidirectional movement of dynein is presented based on the structural observations and biochemical experimental results available. In this model, the binding affinity of dynein for microtubule (MT) is independent of its nucleotide state and the change between strong and weak MT-binding is determined naturally by the variation of relative orientation between the stalk and MT, as the stalk rotates following nucleotide-state transition. Thus the enigmatic communication from the adenosine triphosphate (ATP)-binding site in the globular domain to the far MT-binding site in the tip of the stalk, which is a prerequisite in conventional models, is not required. Using the present model, the previous experimental results such as the effect of ATP and adenosine diphosphate (ADP) bindings on dissociation of dynein from MT, the movement of single-headed axonemal dyneins at saturating ATP concentration, the load dependence of step-size for the movement of two-headed cytoplasmic dyneins and the dependence of stall force on ATP concentration can be well explained.

  4. Monoclonal antibodies in myeloma

    DEFF Research Database (Denmark)

    Sondergeld, P.; van de Donk, N. W. C. J.; Richardson, P. G.;

    2015-01-01

    The development of monoclonal antibodies (mAbs) for the treatment of disease goes back to the vision of Paul Ehrlich in the late 19th century; however, the first successful treatment with a mAb was not until 1982, in a lymphoma patient. In multiple myeloma, mAbs are a very recent and exciting add...

  5. Prediction of Antibody Epitopes

    DEFF Research Database (Denmark)

    Nielsen, Morten; Marcatili, Paolo

    2015-01-01

    self-proteins. Given the sequence or the structure of a protein of interest, several methods exploit such features to predict the residues that are more likely to be recognized by an immunoglobulin.Here, we present two methods (BepiPred and DiscoTope) to predict linear and discontinuous antibody...

  6. Lupus anticoagulants and antiphospholipid antibodies

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/article/000547.htm Lupus anticoagulants and antiphospholipid antibodies To use the sharing features on this page, please enable JavaScript. Lupus anticoagulants are antibodies against substances in the lining ...

  7. Morbillivirus and henipavirus attachment protein cytoplasmic domains differently affect protein expression, fusion support and particle assembly.

    Science.gov (United States)

    Sawatsky, Bevan; Bente, Dennis A; Czub, Markus; von Messling, Veronika

    2016-05-01

    The amino-terminal cytoplasmic domains of paramyxovirus attachment glycoproteins include trafficking signals that influence protein processing and cell surface expression. To characterize the role of the cytoplasmic domain in protein expression, fusion support and particle assembly in more detail, we constructed chimeric Nipah virus (NiV) glycoprotein (G) and canine distemper virus (CDV) haemagglutinin (H) proteins carrying the respective heterologous cytoplasmic domain, as well as a series of mutants with progressive deletions in this domain. CDV H retained fusion function and was normally expressed on the cell surface with a heterologous cytoplasmic domain, while the expression and fusion support of NiV G was dramatically decreased when its cytoplasmic domain was replaced with that of CDV H. The cell surface expression and fusion support functions of CDV H were relatively insensitive to cytoplasmic domain deletions, while short deletions in the corresponding region of NiV G dramatically decreased both. In addition, the first 10 residues of the CDV H cytoplasmic domain strongly influence its incorporation into virus-like particles formed by the CDV matrix (M) protein, while the co-expression of NiV M with NiV G had no significant effect on incorporation of G into particles. The cytoplasmic domains of both the CDV H and NiV G proteins thus contribute differently to the virus life cycle. PMID:26813519

  8. The distribution of special cytoplasmic differentiations of the egg during early cleavage in Limnaea stagnalis

    NARCIS (Netherlands)

    Raven, Chr.P.

    1967-01-01

    Uncleaved eggs of Limnaea stagnalis show a pattern of cytoplasmic differentiations in which there are 6 “subcortical accumulations” (SCA) in the equatorial region. SCA consist of a dense cytoplasmic matrix, containing a special kind of small granules. At certain stages also lens-shaped bodies, consi

  9. Transport of peptidoglycan sub-units across the bacterial cytoplasmic membrane

    NARCIS (Netherlands)

    van Dam, V.

    2009-01-01

    Transport of peptidoglycan sub-units across the bacterial cytoplasmic membrane Vincent van Dam The work described in this thesis focuses on the mechanism by which the lipid-linked cell wall precursor molecule Lipid II is transported from the inner leaflet to the outer leaflet of the cytoplasmic memb

  10. Recombinant antibodies and tumor targeting

    OpenAIRE

    Sheikholvaezin, Ali

    2006-01-01

    Different antibody derived constructs are rapidly advancing as putative tools for treatment of malignant diseases. Antibody engineering has added significant new technologies to modify size, affinities, solubility, stability and biodistribution properties for immunoconjugates. In the present thesis, the aim was to increase our knowledge on how new recombinant antibodies could be tailored to optimize localization to experimental tumors in mice. One hybridoma, producing the monoclonal antibody ...

  11. Regulation of Cytoplasmic and Vacuolar Volumes by Plant Cells in Suspension Culture

    DEFF Research Database (Denmark)

    Owens, Trevor; Poole, Ronald J

    1979-01-01

    Quantitative microscopical measurements have been made of the proportion of cell volume occupied by cytoplasm in a cell suspension culture derived from cotyledons of bush bean (cv. Contender). On a 7-day culture cycle, the content of cytoplasm varies from 25% at the time of transfer to 45% at the...... start of the phase of rapid cell division. If the culture is continued beyond 7 days, the vacuole volume reaches 90% of cell volume by day 12.Attempts to measure relative cytoplasmic volumes by compartmental analysis of nonelectrolyte efflux were unsuccessful. The proportion of cell volume occupied by...... cytoplasm is roughly correlated with protein content, but shows no correlation with cell size or with intracellular concentrations of K or Na. The most striking observation is that the growth of cytoplasmic volume for the culture as a whole appears to be constant throughout the culture cycle, despite...

  12. A cryptic cytoplasmic male sterility unveils a possible gynodioecious past for Arabidopsis thaliana.

    Directory of Open Access Journals (Sweden)

    Nicolas Gobron

    Full Text Available Gynodioecy, the coexistence of hermaphrodites and females (i.e. male-sterile plants in natural plant populations, most often results from polymorphism at genetic loci involved in a particular interaction between the nuclear and cytoplasmic genetic compartments (cytonuclear epistasis: cytoplasmic male sterility (CMS. Although CMS clearly contributes to the coevolution of involved nuclear loci and cytoplasmic genomes in gynodioecious species, the occurrence of CMS genetic factors in the absence of sexual polymorphism (cryptic CMS is not easily detected and rarely taken in consideration. We found cryptic CMS in the model plant Arabidopsis thaliana after crossing distantly related accessions, Sha and Mr-0. Male sterility resulted from an interaction between the Sha cytoplasm and two Mr-0 genomic regions located on chromosome 1 and chromosome 3. Additional accessions with either nuclear sterility maintainers or sterilizing cytoplasms were identified from crosses with either Sha or Mr-0. By comparing two very closely related cytoplasms with different male-sterility inducing abilities, we identified a novel mitochondrial ORF, named orf117Sha, that is most likely the sterilizing factor of the Sha cytoplasm. The presence of orf117Sha was investigated in worldwide natural accessions. It was found mainly associated with a single chlorotype in accessions belonging to a clade predominantly originating from Central Asia. More than one-third of accessions from this clade carried orf117Sha, indicating that the sterilizing-inducing cytoplasm had spread in this lineage. We also report the coexistence of the sterilizing cytoplasm with a non-sterilizing cytoplasm at a small, local scale in a natural population; in addition a correlation between cytotype and nuclear haplotype was detected in this population. Our results suggest that this CMS system induced sexual polymorphism in A. thaliana populations, at the time when the species was mainly outcrossing.

  13. Cytoplasmic p21 is a potential predictor for cisplatin sensitivity in ovarian cancer

    International Nuclear Information System (INIS)

    P21(WAF1/Cip1) binds to cyclin-dependent kinase complexes and inhibits their activities. It was originally described as an inhibitor of cancer cell proliferation. However, many recent studies have shown that p21 promotes tumor progression when accumulated in the cell cytoplasm. So far, little is known about the correlation between cytoplasmic p21 and drug resistance. This study was aimed to investigate the role of p21 in the cisplatin resistance of ovarian cancer. RT-PCR, western blot and immunofluorescence were used to detect p21 expression and location in cisplatin-resistant ovarian cancer cell line C13* and its parental line OV2008. Regulation of cytoplasmic p21 was performed through transfection of p21 siRNA, Akt2 shRNA and Akt2 constitutively active vector in the two cell lines; their effects on cisplatin-induced apoptosis were evaluated by flow cytometry. Tumor tissue sections of clinical samples were analyzed by immunohistochemistry. p21 predominantly localizes to the cytoplasm in C13* compared to OV2008. Persistent exposure to low dose cisplatin in OV2008 leads to p21 translocation from nuclear to cytoplasm, while it had not impact on p21 localization in C13*. Knockdown of cytoplasmic p21 by p21 siRNA transfection in C13* notably increased cisplatin-induced apoptosis through activation of caspase 3. Inhibition of p21 translocation into the cytoplasm by transfection of Akt2 shRNA into C13* cells significantly increased cisplatin-induced apoptosis, while induction of p21 translocation into the cytoplasm by transfection of constitutively active Akt2 in OV2008 enhanced the resistance to cisplatin. Immunohistochemical analysis of clinical ovarian tumor tissues demonstrated that cytoplasmic p21 was negatively correlated with the response to cisplatin based treatment. Cytoplasmic p21 is a novel biomarker of cisplatin resistance and it may represent a potential therapeutic target for ovarian tumors that are refractory to conventional treatment

  14. Antibody mimetics: promising complementary agents to animal-sourced antibodies.

    Science.gov (United States)

    Baloch, Abdul Rasheed; Baloch, Abdul Wahid; Sutton, Brian J; Zhang, Xiaoying

    2016-01-01

    Despite their wide use as therapeutic, diagnostic and detection agents, the limitations of polyclonal and monoclonal antibodies have inspired scientists to design the next generation biomedical agents, so-called antibody mimetics that offer many advantages over conventional antibodies. Antibody mimetics can be constructed by protein-directed evolution or fusion of complementarity-determining regions through intervening framework regions. Substantial progress in exploiting human, butterfly (Pieris brassicae) and bacterial systems to design and select mimetics using display technologies has been made in the past 10 years, and one of these mimetics [Kalbitor® (Dyax)] has made its way to market. Many challenges lie ahead to develop mimetics for various biomedical applications, especially those for which conventional antibodies are ineffective, and this review describes the current characteristics, construction and applications of antibody mimetics compared to animal-sourced antibodies. The possible limitations of mimetics and future perspectives are also discussed. PMID:25264572

  15. A monoclonal antibody against leptin.

    Science.gov (United States)

    Mahmoudian, Jafar; Jeddi-Tehrani, Mahmood; Bayat, Ali Ahmad; Mahmoudi, Ahmad Reza; Vojgani, Yasaman; Tavangar, Banafsheh; Hadavi, Reza; Zarei, Saeed

    2012-10-01

    Leptin is an important protein that regulates energy storage and homeostasis in humans and animals. Leptin deficiency results in various abnormalities such as diabetes, obesity, and infertility. Producing a high affinity monoclonal antibody against human leptin provides an important tool to monitor and trace leptin function in different biological fluids. In this study, recombinant human leptin was conjugated to KLH and injected into mice. After immunization, mouse myeloma SP2/0 cells were fused with murine splenocytes followed by selection of antibody-producing hybridoma cells. After screening of different hybridoma colonies by ELISA, a high affinity antibody was selected and purified by affinity chromatography. The affinity constant of the antibody was measured by ELISA. Western blot, immunocytochemistry, and flow cytometry experiments were used to characterize the antibody. The anti-leptin antibody had a high affinity (around 1.13 × 10(-9) M) for its antigen. The saturation of the antibody with leptin (20 moles leptin per 1 mole antibody) in Western blot analysis proved that the antibody had specific binding to its antigen. Immunocytochemistry and flow cytometry on JEG-3 (human placental choriocarcinoma cell) cells revealed that the anti-leptin antibody recognized intracellular leptin. In conclusion, we report here the production and characterization of a murine anti-leptin antibody with high affinity for human leptin. PMID:23098305

  16. Engineering antibodies by yeast display.

    Science.gov (United States)

    Boder, Eric T; Raeeszadeh-Sarmazdeh, Maryam; Price, J Vincent

    2012-10-15

    Since its first application to antibody engineering 15 years ago, yeast display technology has been developed into a highly potent tool for both affinity maturing lead molecules and isolating novel antibodies and antibody-like species. Robust approaches to the creation of diversity, construction of yeast libraries, and library screening or selection have been elaborated, improving the quality of engineered molecules and certainty of success in an antibody engineering campaign and positioning yeast display as one of the premier antibody engineering technologies currently in use. Here, we summarize the history of antibody engineering by yeast surface display, approaches used in its application, and a number of examples highlighting the utility of this method for antibody engineering.

  17. Antiphospholipid Antibody and Antiphospholipid Syndrome

    Institute of Scientific and Technical Information of China (English)

    吴竞生

    2008-01-01

    @@ Antiphospholipid antibodies (APA) APA is a big category for all kinds of negative charge phospholipid or lecithin - a protein complex autoantibodies or the same antibody, through its recognition of antigen (target protein) different, and phospholipids or lecithin - protein complex combination of various rely on the interference Phospholipid clotting and anti-coagulation factor, and promote endothelial cells, platelets, complement activation and play a role. APA including lupus anticoagulant(LA) and anticardiolipin antibody (ACA), In addition, there are anti-β2 glycoprotein-I (β2-GPI) antibody, anti-prothrombin (a- PT) antibody, anti-lysophosphatidic acid antibody and anti-phosphatidylserine antibody, and so on. APA as the main target of phospholipid-binding protein, including β2-GPI, prothrombin, annexin, protein C (PC) and protein S (PS), plasminogen, and so on.

  18. Intracellular distribution of TM4SF1 and internalization of TM4SF1-antibody complex in vascular endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Sciuto, Tracey E.; Merley, Anne; Lin, Chi-Iou [Center for Vascular Biology Research and Department of Pathology, Beth Israel Deaconess Medical Center and Harvard Medical School (United States); Richardson, Douglas [Department of Molecular and Cellular Biology, Harvard University (United States); Liu, Yu [Department of Pharmacology, Shanxi Medical University, Xinjiannanlu 56, Shanxi Province, Taiyuan 030001 (China); Li, Dan; Dvorak, Ann M. [Center for Vascular Biology Research and Department of Pathology, Beth Israel Deaconess Medical Center and Harvard Medical School (United States); Dvorak, Harold F., E-mail: hdvorak@bidmc.harvard.edu [Center for Vascular Biology Research and Department of Pathology, Beth Israel Deaconess Medical Center and Harvard Medical School (United States); Jaminet, Shou-Ching S., E-mail: sjaminet@bidmc.harvard.edu [Center for Vascular Biology Research and Department of Pathology, Beth Israel Deaconess Medical Center and Harvard Medical School (United States)

    2015-09-25

    Transmembrane-4 L-six family member-1 (TM4SF1) is a small plasma membrane-associated glycoprotein that is highly and selectively expressed on the plasma membranes of tumor cells, cultured endothelial cells, and, in vivo, on tumor-associated endothelium. Immunofluorescence microscopy also demonstrated TM4SF1 in cytoplasm and, tentatively, within nuclei. With monoclonal antibody 8G4, and the finer resolution afforded by immuno-nanogold transmission electron microscopy, we now demonstrate TM4SF1 in uncoated cytoplasmic vesicles, nuclear pores and nucleoplasm. Because of its prominent surface location on tumor cells and tumor-associated endothelium, TM4SF1 has potential as a dual therapeutic target using an antibody drug conjugate (ADC) approach. For ADC to be successful, antibodies reacting with cell surface antigens must be internalized for delivery of associated toxins to intracellular targets. We now report that 8G4 is efficiently taken up into cultured endothelial cells by uncoated vesicles in a dynamin-dependent, clathrin-independent manner. It is then transported along microtubules through the cytoplasm and passes through nuclear pores into the nucleus. These findings validate TM4SF1 as an attractive candidate for cancer therapy with antibody-bound toxins that have the capacity to react with either cytoplasmic or nuclear targets in tumor cells or tumor-associated vascular endothelium. - Highlights: • Anti-TM4SF1 antibody 8G4 was efficiently taken up by cultured endothelial cells. • TM4SF1–8G4 internalization is dynamin-dependent but clathrin-independent. • TM4SF1–8G4 complexes internalize along microtubules to reach the perinuclear region. • Internalized TM4SF1–8G4 complexes pass through nuclear pores into the nucleus. • TM4SF1 is an attractive candidate for ADC cancer therapy.

  19. Cytoplasmic Acidification Induced by Inorganic Phosphate Uptake in Suspension Cultured Catharanthus roseus Cells

    Science.gov (United States)

    Sakano, Katsuhiro; Yazaki, Yoshiaki; Mimura, Tetsuro

    1992-01-01

    Cytoplasmic acidification during inorganic phosphate (Pi) absorption by Catharanthus roseus cells were studied by means of a fluorescent pH indicator, 2′,7′-bis-(2-carboxyethyl)-5 carboxyfluorescein (acetomethylester) (BCECF), and 31P-nuclear magnetic resonance spectroscopy. Cytoplasmic acidification measured by decrease in the fluorescence intensity started immediately after Pi application. Within a minute or so, a stable state was attained and no further acidification occurred, whereas Pi absorption was still proceeding. As soon as Pi in the medium was exhausted, cytoplasmic pH started to recover. Coincidentally, the medium pH started to recover toward the original acidic pH. The Pi-induced changes in the cytoplasmic pH were confirmed by 31P-nuclear magnetic resonance study. Maximum acidification of the cytoplasm induced by 1.7 millimolar Pi was 0.2 pH units. Vacuolar pH was also affected by Pi. In some experiments, but not all, pH decreased reversibly by 0.2 to 0.3 pH units during Pi absorption. Results suggest that the cytoplasmic pH is regulated by proton pumps in the plasma membrane and in the tonoplast. In addition, other mechanisms that could consume extra protons in the cytoplasm are suggested. ImagesFigure 1 PMID:16668939

  20. High Temperature as a Mechanism for Plant Cytoplasm Preservation in Fossils

    Institute of Scientific and Technical Information of China (English)

    WANG Xin

    2007-01-01

    Because the cytoplasm of a plant normally degrades after the death of the plant, finding cytoplasm in a plant body after a prolonged period of time, especially in fossil plants, is unexpected.Recent work on several 100-Myr-old plant fossils from Kansas, USA indicates, however, that cells and their contents can be preserved. Most of the cells in these fossil plants appear to be in a state of plasmolysis, and these fossil cells bear a strong resemblance to laboratory-baked cells of extant plant tissues. Based on a comparison with extant material plus biophysical and biochemical analyses of the cytoplasm degrading process, a new hypothesis for cytoplasm preservation in nature is proposed: high temperature, a concomitant of commonly seen wildfires, may preserve cytoplasm in fossil plants. This hypothesis implies that fossilized cytoplasm should be rather common and an appropriate substance for research, unlike previously thought. Research on fossil cytoplasm closely integrates paleobotany with biochemistry, biophysics, as well as fire ecology, and invites inputs from these fields to paleobotany to interpret these provocative findings.

  1. Genetics, Development, and Application of Cytoplasmic Herbicide Resistance in Foxtail Millet

    Institute of Scientific and Technical Information of China (English)

    JI Gui-su; DU Rui-heng; HOU Sheng-lin; CHENG Ru-hong; WANG Xin-yu; ZHAO Xiu-ping

    2007-01-01

    The effect of cytoplasmic herbicide resistant gene in millet plants was studied. The heterozygous populations and isogenic lines with homocaryotic alloplasmic genes were obtained by crossing and reciprocal crossing of cytoplasmic herbicide resistant plants with susceptive plants of foxtail millet. The characters of F1, F2, backcross and composite cross groups, and the growth and development of isogenic lines were compared. The cytoplasmic herbicide resistant gene slowed the development of seedling, delayed heading, and shortened the milking stage in the foxtail millet plant. Yield capacity and main agronomic characters were all affected by the cytoplasmic herbicide resistant gene in most of the backcross, composite cross, and F2 populations. However, there was stronger hybrid vigor in F1. The backcrosses,composite crosses, and F2 populations were widely separated and some of them had good characters similar to those of susceptive groups. The plant characters and development of foxtail millet were negatively affected by the cytoplasmic herbicide resistant gene. The authors proposed a method of using hybrid vigor to obtain high yield and avoid the negative effects of herbicide resistance cytoplasm in plant growth. The expected results could be obtained by selecting individuals in separate populations of fast developed seedlings, well-developed roots, and with capacities of early heading and fast milking. Guided by the principal mentioned above, many high yield lines and hybrid crosses of foxtail millet with herbicide resistant cytoplasm were obtained.

  2. Suppurative dacroadenitis causing ocular sicca syndrome in classic Wegener′s granulomatosis

    Directory of Open Access Journals (Sweden)

    Khanna Dhanita

    2011-01-01

    Full Text Available Wegener′s granulomatosis (WG is a multisystem vasculitic disorder which can commonly afflict various components of the eye. Here we describe some unusual ocular manifestations of the disease in one patient. A young male with history of upper respiratory tract symptoms including epistaxis, nasal stuffiness and maxillary sinus pain presented with bilateral lacrimal gland abscess and ptosis. Lacrimal gland biopsy revealed granulomatous vasculitis. Lung cavities, positive cytoplasmic-antineutrophil cytoplasmic antibodies and high titers of serine proteinase-3 antibodies confirmed the diagnosis of WG. The patient developed dry eyes after a month of first presentation. There was no dryness of mouth, suggesting the absence of salivary gland involvement, and antinuclear antibodies as well as antibodies against Ro and La antigens classical of primary Sjogren′s syndrome were absent. Granulomatous vasculitis of lacrimal gland leading to abscess formation and dryness of eyes has not been described in WG and reflects the aggressive nature of inflammatory process in this disease.

  3. Detection of serum anti-melanocyte antibodies and identification of related antigens in patients with vitiligo.

    Science.gov (United States)

    Zhu, M C; Liu, C G; Wang, D X; Zhan, Z

    2015-12-07

    We detected autoantibodies against melanocytes in serum samples obtained from 50 patients, including 4 with HBV, with vitiligo and identified the associated membrane antigens. Heat shock protein 70 (HSP70) and anti-tyrosinase-related protein 1 (TRP-1) antibody levels were analyzed. The associated antigens in normal human melanocyte were identified by immunofluorescence. Autoantibodies against melanocyte membrane and cytoplasmic proteins were detected by western blot. Membrane antigens with higher frequencies were identified by protein mass spectrometry. The HSP70 and anti-TRP-1 antibody levels (N = 70; 10 with HBV) were detected by ELISA. The specific antigens were detected in melanocyte cytoplasm and membrane (40/50; 80% incidence; western blot). The autoantibodies reacted with several membrane antigens with approximate molecular weights (Mr) of 86,000, 75,000, 60,000, 52,000, and 44,000 (strip positive rates: 36, 58, 22, 2, and 2%, respectively). Thirty percent of the patients showed the presence of cytoplasmic antigens (Mr: 110,000, 90,000, 75,000, 50,000, and 400,000; strip positive rates: 12, 4, 12, 10, and 2%, respectively). Fifteen and 5% of the healthy subjects showed positive expression of membrane and cytoplasmic antigens, respectively. Protein mass spectrometry predicted membrane proteins with Mr of 86,000 and 75,000 and 60,000 to be Lamin A /C and Vimentin X1, respective. High titers of anti-TRP-1 antibody were detected and showed positive correlation with HSP70 (r = 0. 927, P vitiligo, which might assist future investigations into autoimmune pathogenesis of vitiligo and formation of autoantibodies. HBV infection was correlated to vitiligo.

  4. Measurement of Cytoplasmic Streaming in Chara Corallina by Magnetic Resonance Velocimetry

    CERN Document Server

    van de Meent, Jan-Willem; Gladden, Lynn F; Goldstein, Raymond E

    2009-01-01

    In aquatic plants such as the Characean algae, the force generation that drives cyclosis is localized within the cytoplasm, yet produces fluid flows throughout the vacuole. For this to occur the tonoplast must transmit hydrodynamic shear efficiently. Here, using magnetic resonance velocimetry, we present the first whole-cell measurements of the cross-sectional longitudinal velocity field in Chara corallina and show that it is in quantitative agreement with a recent theoretical analysis of rotational cytoplasmic streaming driven by bidirectional helical forcing in the cytoplasm, with direct shear transmission by the tonoplast.

  5. Primary observations of the existence of Fas-like cytoplasmic death factor in plant cells

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    The main activity of Fas is to trigger cytoplasm death program in animal cells. In G2 pea, vacuole plays a pivotal role in inducing cell death in the cytoplasm of longday (LD) grown apical meristem cells. Expression patterns of the Fas in G2 pea cells revealed that the Fas is mainly localized in the vacuole of cells undergoing programmed cell death (PCD). The Fas expression is corresponding to the initiation of menadione-induced PCD in tobacco protoplasts.The results suggest the existence of the Fas-like mediated cytoplasmic death pathway in plant cells.``

  6. Transcytosis of Aminopeptidase N in caco-2 cells is mediated by a Non-cytoplasmic Signal

    DEFF Research Database (Denmark)

    Vogel, L K; Norén, Ove; Sjöström, H

    1995-01-01

    transmembrane or cytoplasmic domain of aminopeptidase N for transport of aminopeptidase N by the indirect pathway by analysis of mutated forms of aminopeptidase N recombinantly expressed in Caco-2 cells. A tail-less and two secretory forms of aminopeptidase N, all deprived of the cytoplasmic tail, were...... transported to the basolateral plasma membrane in proportions equivalent to the wild type enzyme. This shows that no cytoplasmic basolateral sorting signal is involved in directing aminopeptidase N to the basolateral plasma membrane. Both the wild type and the tail-less aminopeptidase N were transcytosed from...

  7. ACE inhibitors can induce circulating antibodies directed to antigens of the superficial epidermal cells.

    Science.gov (United States)

    Cozzani, Emanuele; Rosa, Gian Marco; Drosera, Massimo; Intra, Chiara; Barsotti, Antonio; Parodi, Aurora

    2011-07-01

    Drug-induced pemphigus has been reported in patients receiving angiotensin-converting enzyme inhibitors. The aim of this work was to study a group of hypertensive patients without skin diseases treated with angiotensin-converting enzyme (ACE) Inhibitors (I), to verify the presence of serum circulating anti-antibodies. The indirect immunofluorescence showed that 33 sera (52.38%) presented autoantibodies directed to an antigen of the cytoplasm of the superficial epidermal keratinocytes. Two of the 33 positive sera had antibodies to Dsg1 and/or 3 in ELISA. Immunoblot analyses were negative. All the 48 control sera were found to have no circulating antibodies using the three assays. Our results would confirm that ACEI drugs may trigger the production of circulating autoantibodies also in patients without clinical manifestations of pemphigus. PMID:20563876

  8. Development of novel monoclonal antibodies that define differentiation stages of human stromal (mesenchymal) stem cells

    DEFF Research Database (Denmark)

    Andersen, Ditte Caroline; Kortesidis, Angela; Zannettino, Andrew C W;

    2011-01-01

    Human mesenchymal stem cells (hMSC) are currently being introduced for cell therapy, yet, antibodies specific for native and differentiated MSCs are required for their identification prior to clinical use. Herein, high quality antibodies against MSC surface proteins were developed by immunizing...... differentiation. Interestingly, undifferentiated cells revealed a sole cytoplasmic distribution pattern of Collagen VI, which however changed to an extracellular matrix appearance upon osteogenic- and adipogenic differentiation. In relation to this, we found that STRO-1(+/-)/Collagen VI(-) sorted hMSC contained...... fewer differentiated alkaline phosphatase(+) cells compared to STRO-1(+/-)/Collagen VI(+) hMSC, suggesting that Collagen VI on the cell membrane exclusively defines differentiated MSCs. In conclusion, we have generated a panel of high quality antibodies to be used for characterization of MSCs, and in...

  9. The antibody Hijikata Tatsumi

    Directory of Open Access Journals (Sweden)

    Éden Peretta

    2012-11-01

    Full Text Available Considered one of the most influential modern dance representatives in Japan, Tatsumi Hijikata’s work was a milestone in the Japanese post-war experimental artistic scene. Heretic son of his time, he staged a fertile mix of artistic and cultural influences, overlapping subversive elements of European arts and philosophy with radical references from pre-modern Japanese culture. In this way he built the foundations of its unstable antibody, its political-artistic project of dissolution of a organism, both physical and social.

  10. Evolutionary genomics of nucleo-cytoplasmic large DNA viruses.

    Science.gov (United States)

    Iyer, Lakshminarayan M; Balaji, S; Koonin, Eugene V; Aravind, L

    2006-04-01

    A previous comparative-genomic study of large nuclear and cytoplasmic DNA viruses (NCLDVs) of eukaryotes revealed the monophyletic origin of four viral families: poxviruses, asfarviruses, iridoviruses, and phycodnaviruses [Iyer, L.M., Aravind, L., Koonin, E.V., 2001. Common origin of four diverse families of large eukaryotic DNA viruses. J. Virol. 75 (23), 11720-11734]. Here we update this analysis by including the recently sequenced giant genome of the mimiviruses and several additional genomes of iridoviruses, phycodnaviruses, and poxviruses. The parsimonious reconstruction of the gene complement of the ancestral NCLDV shows that it was a complex virus with at least 41 genes that encoded the replication machinery, up to four RNA polymerase subunits, at least three transcription factors, capping and polyadenylation enzymes, the DNA packaging apparatus, and structural components of an icosahedral capsid and the viral membrane. The phylogeny of the NCLDVs is reconstructed by cladistic analysis of the viral gene complements, and it is shown that the two principal lineages of NCLDVs are comprised of poxviruses grouped with asfarviruses and iridoviruses grouped with phycodnaviruses-mimiviruses. The phycodna-mimivirus grouping was strongly supported by several derived shared characters, which seemed to rule out the previously suggested basal position of the mimivirus [Raoult, D., Audic, S., Robert, C., Abergel, C., Renesto, P., Ogata, H., La Scola, B., Suzan, M., Claverie, J.M. 2004. The 1.2-megabase genome sequence of Mimivirus. Science 306 (5700), 1344-1350]. These results indicate that the divergence of the major NCLDV families occurred at an early stage of evolution, prior to the divergence of the major eukaryotic lineages. It is shown that subsequent evolution of the NCLDV genomes involved lineage-specific expansion of paralogous gene families and acquisition of numerous genes via horizontal gene transfer from the eukaryotic hosts, other viruses, and bacteria

  11. VIRAL ANTIBODIES IN PRESCHOOL CHILDREN

    Directory of Open Access Journals (Sweden)

    S. Saidi

    1974-08-01

    Full Text Available One hundred sera from children 1 - 6 years of age, representative of a large serum collection, were tested for the prevalence of antibodies against different viruses. Hemagglutination-inhibition (HI antibodies were found in 68% for measles; 61 % for rubella; 75'% for influenza A2/Hong Kong/68, 16% for influenza B/Md./59, 0% for group A arboviruses, 10% for group B arboviruses, 3% for phlebotomus fever group and 4% for Congo-Crimean hemorrhagic fever (C-CHF group of arboviruses Poliomyelitis-neutralizing antibodies for type 1, 2 and 3 were 90%; 85% and 84%~ respectively. Antibody to EH virus was detected in 84% of the sera by immuno-fluorescence. None of the sera were positive for hepatitis-B antigen or antibody by immuno-precipitation test. The prevalence of some viral antibodies found in this survey are compared with results obtained from surveys in other parts of the country.

  12. Organization of the cytoplasmic reticulum in the central vacuole of parenchyma cells in Allium cepa L.

    Directory of Open Access Journals (Sweden)

    Tomasz J. Wodzicki

    2015-05-01

    Full Text Available An elaborate and complex cytoplasmic reticulum composed of fine filaments and lamellae ranging from 0.1 to 4 microns in size is revealed by viewing the central vacuole of onion bulb parenchyma cells with the scanning election microscope. The larger cytoplasmic strands, visible with the light microscope, are composed of numerous smaller filaments (some tubular which might explain the observed bidirectional movement of particles in these larger strands. The finely divided cytoplasmic network of filaments is continuous with the parietal cytoplasm inclosing the vacuolar sap. In these highly vacuolated cells the mass of the protoplast is in the form of an intravacuolar reticulum immersed in the cell sap. The probable significance of the vacuolar sap in relation to physiological processes of the cell is discussed.

  13. Reactive oxygen species (ROS) is not a promotor of taxol-induced cytoplasmic vacuolization

    Science.gov (United States)

    Sun, Qingrui; Chen, Tongsheng

    2009-02-01

    we have previously reported that taxol, a potent anticancer agent, induces caspase-independent cell death and cytoplasmic vacuolization in human lung adenocarcinoma (ASTC-a-1) cells. However, the mechanisms of taxol-induced cytoplasmic vacuolization are poorly understood. Reactive oxygen species (ROS) has been reported to be involved in the taxol-induced cell death. Here, we employed confocal fluorescence microscopy imaging to explore the role of ROS in taxol-induced cytoplasmic vacuolization. We found that ROS inhibition by addition of N-acetycysteine (NAC), a total ROS scavenger, did not suppress these vacuolization but instead increased vacuolization. Take together, our results showed that ROS is not a promotor of the taxol-induced cytoplasmic vacuolization.

  14. PRELIMINARY RESULTS OF INTRA CYTOPLASMIC SPERM INJECTION (ICSI) APPLIED ON RABBIT : A TECHNICAL NOTE.

    OpenAIRE

    Escribá, M.J.; García- Ximénez, F.

    1999-01-01

    Abstract not available. Escribá, M.; García- Ximénez, F. (1999). PRELIMINARY RESULTS OF INTRA CYTOPLASMIC SPERM INJECTION (ICSI) APPLIED ON RABBIT : A TECHNICAL NOTE. http://hdl.handle.net/10251/10364.

  15. Cytoplasm enhancement operator of peripheral blood smear images that are instable-stained and overexposed

    Science.gov (United States)

    Zheng, Xin; Wang, Guoyou; Liu, Jianguo

    2015-12-01

    Nucleus and cytoplasm are both essential for white blood cell recognition but the edges of cytoplasm are too blurry to be detected because of instable staining and overexposure. This paper aims at proposing a cytoplasm enhancement operator (CEO) to achieve accurate convergence of the active contour model. The CEO contains two parts. First, a nonlinear over-exposure enhancer map is yielded to correct over-exposure, which suppresses background noise while preserving details and improving contrast. Second, the over-exposed regions of cytoplasm in particular is further enhanced by a tri- modal histogram specification based on the scale-space filtering. The experimental results show that the proposed CEO and its corresponding GVF snake is superior to other unsupervised segmentation approaches.

  16. The nucleotide sequences of the initiator transfer RNAs from bean cytoplasm and chloroplasts.

    OpenAIRE

    Canaday, J; Guillemaut, P; Weil, J H

    1980-01-01

    The initiator tRNAsMet from the cytoplasm and chloroplasts of Phaseolus vulgaris have been purified and sequenced. The sequence of bean cytoplasmic initiator tRNAiMet is : pA-U-C-A-G-A-G-U-m1G-m2G-C-G-C-A-G-C-G-G-A-A-G-C-G-U-m2G-G-U-G-G-G2-C-C-C-A-U-t6A-A-C-C-C-A-C-A-G-m7G-D-m5C-C-C-A-G-G-A-psi-C-G-m1A-A-A-C-C-U-Gm-G-C-U-C-U-G-A-U-A-C-C-AOH. The sequence of bean cytoplasmic tRNAiMet is almost identical to that of wheat germ and shows a high degree of homology with other cytoplasmic initiator ...

  17. Nonlinearity in cytoplasm viscosity can generate an essential symmetry breaking in cellular behaviors.

    Science.gov (United States)

    Tachikawa, Masashi; Mochizuki, Atsushi

    2015-01-01

    The cytoplasms of ameboid cells are nonlinearly viscous. The cell controls this viscosity by modulating the amount, localization and interactions of bio-polymers. Here we investigated how the nonlinearity infers the cellular behaviors and whether nonlinearity-specific behaviors exist. We modeled the developed plasmodium of the slime mold Physarum polycephalum as a network of branching tubes and examined the linear and nonlinear viscous cytoplasm flows in the tubes. We found that the nonlinearity in the cytoplasm׳s viscosity induces a novel type of symmetry breaking in the protoplasmic flow. We also show that symmetry breaking can play an important role in adaptive behaviors, namely, connection of behavioral modes implemented on different time scales and transportation of molecular signals from the front to the rear of the cell during cellular locomotion. PMID:25261729

  18. Novel nuclear-cytoplasmic interaction in wheat (Triticum aestivum) induces vigorous plants

    Science.gov (United States)

    Interspecific hybridization can be considered an accelerator of evolution, otherwise a slow process, solely dependent on mutation and recombination. Upon interspecific hybridization, several novel interactions between nuclear and cytoplasmic genomes emerge which provide additional sources of diversi...

  19. Secretion of Cpn0796 from Chlamydia pneumoniae into the host cell cytoplasm by an autotransporter mechanism

    DEFF Research Database (Denmark)

    Vandahl, Brian B S; Stensballe, Allan; Roepstorff, Peter;

    2005-01-01

    infected cells, whereas only the 65 kDa full-length Cpn0796 could be detected in purified Chlamydia. Confocal immunofluorescence microscopy demonstrated that Cpn0796 was localized in the Chlamydia membrane in young inclusions. However, at 36 h post infection and later Cpn0796 was detected in the cytoplasm...... of C. pneumoniae infected HEp-2 and BHK cells. Furthermore, Cpn0796 was detected in the cytoplasm of infected cells in the lungs of C. pneumoniae infected C57Bl mice. When cleavage was inhibited, Cpn0796 was retained in the chlamydiae. We propose that Cpn0796 is an autotransporter the N-terminal of...... which is translocated to the host cell cytoplasm. This is the first example of secretion of a Chlamydia autotransporter passenger domain into the host cell cytoplasm. Cpn0796 is specific for C. pneumoniae, where five homologous proteins are encoded by clustered genes. None of these five proteins were...

  20. Metrics for antibody therapeutics development

    OpenAIRE

    Reichert, Janice M

    2010-01-01

    A wide variety of full-size monoclonal antibodies (mAbs) and therapeutics derived from alternative antibody formats can be produced through genetic and biological engineering techniques. These molecules are now filling the preclinical and clinical pipelines of every major pharmaceutical company and many biotechnology firms. Metrics for the development of antibody therapeutics, including averages for the number of candidates entering clinical study and development phase lengths for mAbs approv...

  1. Empowered Antibody Therapies - IBC conference.

    Science.gov (United States)

    Herold, Jens

    2010-10-01

    The Empowered Antibody Therapies conference, held in Burlingame, CA, USA, included topics covering new therapeutic developments in the field of multispecific antibodies. This conference report highlights selected presentations on DVD-Igs from Abbott Laboratories, ImmTACs from Immunocore, 'Dock-and-Lock' technology from Immunomedics, the bispecific BiTE antibody blinatumomab from Micromet, and Triomabs from TRION Pharma and Fresenius Biotech. PMID:20878591

  2. Monoclonal antibodies for treating cancer

    International Nuclear Information System (INIS)

    The purpose of this study is to assess the current status of in-vivo use of monoclonal antibodies for treating cancer. Publications appearing between 1980 and 1988 were identified by computer searches using MEDLINE and CANCERLIT, by reviewing the table of contents of recently published journals, and by searching bibliographies of identified books and articles. More than 700 articles, including peer-reviewed articles and book chapters, were identified and selected for analysis. The literature was reviewed and 235 articles were selected as relevant and representative of the current issues and future applications for in-vivo monoclonal antibodies for cancer therapy and of the toxicity and efficacy which has been associated with clinical trials. Approaches include using antibody alone (interacting with complement or effector cells or binding directly with certain cell receptors) and immunoconjugates (antibody coupled to radioisotopes, drugs, toxins, or other biologicals). Most experience has been with murine antibodies. Trials of antibody alone and radiolabeled antibodies have confirmed the feasibility of this approach and the in-vivo trafficking of antibodies to tumor cells. However, tumor cell heterogeneity, lack of cytotoxicity, and the development of human antimouse antibodies have limited clinical efficacy. Although the immunoconjugates are very promising, heterogeneity and the antimouse immune response have hampered this approach as has the additional challenge of chemically or genetically coupling antibody to cytotoxic agents. As a therapeutic modality, monoclonal antibodies are still promising but their general use will be delayed for several years. New approaches using human antibodies and reducing the human antiglobulin response should facilitate treatment. 235 references

  3. Cytoplasmic long noncoding RNAs are frequently bound to and degraded at ribosomes in human cells

    OpenAIRE

    Carlevaro-Fita, Joana; Rahim, Anisa; Guigó, Roderic; Vardy, Leah A.; Johnson, Rory

    2016-01-01

    Recent footprinting studies have made the surprising observation that long noncoding RNAs (lncRNAs) physically interact with ribosomes. However, these findings remain controversial, and the overall proportion of cytoplasmic lncRNAs involved is unknown. Here we make a global, absolute estimate of the cytoplasmic and ribosome-associated population of stringently filtered lncRNAs in a human cell line using polysome profiling coupled to spike-in normalized microarray analysis. Fifty-four percent ...

  4. Secretion of Bacterial Lipoproteins: Through the Cytoplasmic Membrane, the Periplasm and Beyond

    OpenAIRE

    Zückert, Wolfram R.

    2014-01-01

    Bacterial lipoproteins are peripherally anchored membrane proteins that play a variety of roles in bacterial physiology and virulence in monoderm (single membrane-enveloped, e.g., grampositive) and diderm (double membrane-enveloped, e.g., gram-negative) bacteria. After export of prolipoproteins through the cytoplasmic membrane, which occurs predominantly but not exclusively via the general secretory or Sec pathway, the proteins are lipid-modified at the cytoplasmic membrane in a multistep pro...

  5. The genetic and molecular basis of cytoplasmic male sterility and fertility restoration in rice

    Institute of Scientific and Technical Information of China (English)

    GUO JingXin; LIU YaoGuang

    2009-01-01

    Cytoplasmic male sterility (CMS) is a maternally inherited characteristic found in many (>150) plant species. CMS/restoration systems are useful tools for hybrid seed production, and are ideal models for study of the interactions between nuclear and mitochondrial genomes. CMS/restoration systems in rice have been widely used for hybrid seed production, greatly contributing to the food supply. This article reviews the progress of the studies on the genetic and molecular basis of cytoplasmic male sterility and fertility restoration in rice.

  6. The entry of diphtheria toxin into the mammalian cell cytoplasm: evidence for lysosomal involvement

    OpenAIRE

    1980-01-01

    Lysosomotropic amines, such as ammonium chloride, are known to protect cells from the cytotoxic effects of diphtheria toxin. These drugs are believed to inhibit the transport of the toxin from a receptor at the cell exterior into the cytoplasm where a fragment of the toxin arrests protein synthesis. We studied the effects of lysosomotropic agents on the cytotoxic process to better understand how the toxin enters the cytoplasm. The cytotoxic effects of diphtheria toxin were not inhibited by an...

  7. Microtubule-microtubule sliding by kinesin-1 is essential for normal cytoplasmic streaming in Drosophila oocytes.

    Science.gov (United States)

    Lu, Wen; Winding, Michael; Lakonishok, Margot; Wildonger, Jill; Gelfand, Vladimir I

    2016-08-23

    Cytoplasmic streaming in Drosophila oocytes is a microtubule-based bulk cytoplasmic movement. Streaming efficiently circulates and localizes mRNAs and proteins deposited by the nurse cells across the oocyte. This movement is driven by kinesin-1, a major microtubule motor. Recently, we have shown that kinesin-1 heavy chain (KHC) can transport one microtubule on another microtubule, thus driving microtubule-microtubule sliding in multiple cell types. To study the role of microtubule sliding in oocyte cytoplasmic streaming, we used a Khc mutant that is deficient in microtubule sliding but able to transport a majority of cargoes. We demonstrated that streaming is reduced by genomic replacement of wild-type Khc with this sliding-deficient mutant. Streaming can be fully rescued by wild-type KHC and partially rescued by a chimeric motor that cannot move organelles but is active in microtubule sliding. Consistent with these data, we identified two populations of microtubules in fast-streaming oocytes: a network of stable microtubules anchored to the actin cortex and free cytoplasmic microtubules that moved in the ooplasm. We further demonstrated that the reduced streaming in sliding-deficient oocytes resulted in posterior determination defects. Together, we propose that kinesin-1 slides free cytoplasmic microtubules against cortically immobilized microtubules, generating forces that contribute to cytoplasmic streaming and are essential for the refinement of posterior determinants.

  8. Cytoplasmic long noncoding RNAs are frequently bound to and degraded at ribosomes in human cells.

    Science.gov (United States)

    Carlevaro-Fita, Joana; Rahim, Anisa; Guigó, Roderic; Vardy, Leah A; Johnson, Rory

    2016-06-01

    Recent footprinting studies have made the surprising observation that long noncoding RNAs (lncRNAs) physically interact with ribosomes. However, these findings remain controversial, and the overall proportion of cytoplasmic lncRNAs involved is unknown. Here we make a global, absolute estimate of the cytoplasmic and ribosome-associated population of stringently filtered lncRNAs in a human cell line using polysome profiling coupled to spike-in normalized microarray analysis. Fifty-four percent of expressed lncRNAs are detected in the cytoplasm. The majority of these (70%) have >50% of their cytoplasmic copies associated with polysomal fractions. These interactions are lost upon disruption of ribosomes by puromycin. Polysomal lncRNAs are distinguished by a number of 5' mRNA-like features, including capping and 5'UTR length. On the other hand, nonpolysomal "free cytoplasmic" lncRNAs have more conserved promoters and a wider range of expression across cell types. Exons of polysomal lncRNAs are depleted of endogenous retroviral insertions, suggesting a role for repetitive elements in lncRNA localization. Finally, we show that blocking of ribosomal elongation results in stabilization of many associated lncRNAs. Together these findings suggest that the ribosome is the default destination for the majority of cytoplasmic long noncoding RNAs and may play a role in their degradation.

  9. Structure of bacterial cytoplasmic chemoreceptor arrays and implications for chemotactic signaling.

    Science.gov (United States)

    Briegel, Ariane; Ladinsky, Mark S; Oikonomou, Catherine; Jones, Christopher W; Harris, Michael J; Fowler, Daniel J; Chang, Yi-Wei; Thompson, Lynmarie K; Armitage, Judith P; Jensen, Grant J

    2014-01-01

    Most motile bacteria sense and respond to their environment through a transmembrane chemoreceptor array whose structure and function have been well-studied, but many species also contain an additional cluster of chemoreceptors in their cytoplasm. Although the cytoplasmic cluster is essential for normal chemotaxis in some organisms, its structure and function remain unknown. Here we use electron cryotomography to image the cytoplasmic chemoreceptor cluster in Rhodobacter sphaeroides and Vibrio cholerae. We show that just like transmembrane arrays, cytoplasmic clusters contain trimers-of-receptor-dimers organized in 12-nm hexagonal arrays. In contrast to transmembrane arrays, however, cytoplasmic clusters comprise two CheA/CheW baseplates sandwiching two opposed receptor arrays. We further show that cytoplasmic fragments of normally transmembrane E. coli chemoreceptors form similar sandwiched structures in the presence of molecular crowding agents. Together these results suggest that the 12-nm hexagonal architecture is fundamentally important and that sandwiching and crowding can replace the stabilizing effect of the membrane. DOI: http://dx.doi.org/10.7554/eLife.02151.001.

  10. The Composition and Organization of Cytoplasm in Prebiotic Cells

    Directory of Open Access Journals (Sweden)

    Jack T. Trevors

    2011-03-01

    Full Text Available This article discusses the hypothesized composition and organization of cytoplasm in prebiotic cells from a theoretical perspective and also based upon what is currently known about bacterial cytoplasm. It is unknown if the first prebiotic, microscopic scale, cytoplasm was initially contained within a primitive, continuous, semipermeable membrane, or was an uncontained gel substance, that later became enclosed by a continuous membrane. Another possibility is that the first cytoplasm in prebiotic cells and a primitive membrane organized at the same time, permitting a rapid transition to the first cell(s capable of growth and division, thus assisting with the emergence of life on Earth less than a billion years after the formation of the Earth. It is hypothesized that the organization and composition of cytoplasm progressed initially from an unstructured, microscopic hydrogel to a more complex cytoplasm, that may have been in the volume magnitude of about 0.1–0.2 µm3 (possibly less if a nanocell prior to the first cell division.

  11. Exploration of cytoplasmic inheritance as a contributor to maternal effects in Welsh Mountain sheep.

    Science.gov (United States)

    Pritchard, Tracey; Cahalan, Christine; Ap Dewi, Ioan

    2008-01-01

    Cytoplasmic effects were investigated using a dataset comprising three breeding groups of Welsh Mountain sheep. The influences of cytoplasmic effects were investigated by comparing animal models with and without a random term representing cytoplasmic effects. The models were applied to the eight-week weight, scan weight (mean 152 days) and ultrasonically scanned muscle and fat depth. The animal model included the random effects of animals and the maternal additive genetic, maternal permanent environmental and maternal common environmental effects. In total there were 24 569, 10 509, 8389, 8369 records for the eight-week weight, scan weight, muscle depth and fat depth respectively. Four subsets were further analysed containing maternal lines with at least five, ten, fifteen and twenty animals/line. There was no evidence of cytoplasmic effects on eight-week weight and muscle depth. Cytoplasmic effects contributed 1-2% of phenotypic variance for scan-weight and fat depth, but the effect was generally non-significant (P >0.05). As the number of animals per maternal line increased, the magnitude of cytoplasmic effects also increased for these traits. Direct heritability estimates for the eight-week weight, scan weight, muscle depth and fat depth using the full dataset were 0.18, 0.25, 0.24, and 0.21 respectively.

  12. Exploration of cytoplasmic inheritance as a contributor to maternal effects in Welsh Mountain sheep

    Directory of Open Access Journals (Sweden)

    Dewi Ioan

    2008-05-01

    Full Text Available Abstract Cytoplasmic effects were investigated using a dataset comprising three breeding groups of Welsh Mountain sheep. The influences of cytoplasmic effects were investigated by comparing animal models with and without a random term representing cytoplasmic effects. The models were applied to the eight-week weight, scan weight (mean 152 days and ultrasonically scanned muscle and fat depth. The animal model included the random effects of animals and the maternal additive genetic, maternal permanent environmental and maternal common environmental effects. In total there were 24 569, 10 509, 8389, 8369 records for the eight-week weight, scan weight, muscle depth and fat depth respectively. Four subsets were further analysed containing maternal lines with at least five, ten, fifteen and twenty animals/line. There was no evidence of cytoplasmic effects on eight-week weight and muscle depth. Cytoplasmic effects contributed 1–2% of phenotypic variance for scan-weight and fat depth, but the effect was generally non-significant (P > 0.05. As the number of animals per maternal line increased, the magnitude of cytoplasmic effects also increased for these traits. Direct heritability estimates for the eight-week weight, scan weight, muscle depth and fat depth using the full dataset were 0.18, 0.25, 0.24, and 0.21 respectively.

  13. Localization and function of KLF4 in cytoplasm of vascular smooth muscle cell

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Yan [Department of Biochemistry and Molecular Biology, The Key Laboratory of Neurobiology and Vascular Biology (China); The Third Hospital of Hebei Medical University, Shijazhuang (China); Zheng, Bin; Zhang, Xin-hua; Nie, Chan-juan; Li, Yong-hui [Department of Biochemistry and Molecular Biology, The Key Laboratory of Neurobiology and Vascular Biology (China); Wen, Jin-kun, E-mail: wjk@hebmu.edu.cn [Department of Biochemistry and Molecular Biology, The Key Laboratory of Neurobiology and Vascular Biology (China)

    2013-06-28

    Highlights: •PDGF-BB prompts the translocation of KLF4 to the cytoplasm. •PDGF-BB promotes interaction between KLF4 and actin in the cytoplasm. •Phosphorylation and SUMOylation of KLF4 participates in regulation of cytoskeletal organization. •KLF4 regulates cytoskeleton by promoting the expression of contraction-associated genes. -- Abstract: The Krüppel-like factor 4 is a DNA-binding transcriptional regulator that regulates a diverse array of cellular processes, including development, differentiation, proliferation, and apoptosis. The previous studies about KLF4 functions mainly focused on its role as a transcription factor, its functions in the cytoplasm are still unknown. In this study, we found that PDGF-BB could prompt the translocation of KLF4 to the cytoplasm through CRM1-mediated nuclear export pathway in vascular smooth muscle cells (VSMCs) and increased the interaction of KLF4 with actin in the cytoplasm. Further study showed that both KLF4 phosphorylation and SUMOylation induced by PDGF-BB participates in regulation of cytoskeletal organization by stabilizing the actin cytoskeleton in VSMCs. In conclusion, these results identify that KLF4 participates in the cytoskeletal organization by stabilizing cytoskeleton in the cytoplasm of VSMCs.

  14. Pathogenesis of Focal Cytoplasmic Necrosis of the Smooth Muscle Cells in Hypertensive Rat Arterial Media

    International Nuclear Information System (INIS)

    Hypertensive rat arteries exhibited severe medial smooth muscle cell injury and necrosis. Electron microscopic observations showed the smooth muscle cells of these arteries exhibited characteristics of focal cytoplasmic necrosis forming new cytodemarcating membrane between the healthy cytoplasm and necrotic cytoplasm. When the focal necrotic cytoplasm disappeared from the injured smooth muscle cells, it left it with a moth-eaten leaf-like appearance (moth-eaten necrosis). At an advanced stage of injury, smooth muscle cells changed to islet-like cell bodies with newly formed basement membranes around them, and further islet-like cell bodies and cell debris disappeared leaving lamellar and reticular basement membranes. In hypertensive rats injected with nitroblue tetrazolium (NBT), formazan deposits were observed in the medial cells and nitrotyrosine, a biomarker of peroxynitrite, were immunohistochemically observed in the arterial media. Nick-end positive extranuclear small granular bodies, which might have derived from focal necrotic cytoplasm and nucleus, were detected in the arterial media using DNA nick-end labeling method. Based on electron microscopical and histochemical findings, we conjectured that the focal cytoplasmic necrosis of the smooth muscle cells in the arterial media depended on injury arising from mitochondria-derived oxidants

  15. Evaluation of line and breed of cytoplasm effects on performance of purebred Brangus cattle.

    Science.gov (United States)

    Rohrer, G A; Taylor, J F; Sanders, J O; Thallman, R M

    1994-11-01

    Substantial differences between reciprocally crossed Bos taurus x Bos indicus calves for birth, weaning, and yearling weights have been reported. To determine whether cytoplasmic inheritance is responsible for a portion of these differences, field records for birth and weaning weight (n = 7,353) and postweaning average daily gain (n = 2,746) from registered Brangus calves were analyzed. An animal model that included maternal effects was fit for each trait. Breed of cytoplasmic origin was fit as a fixed effect and coded as Angus, Brahman, or unknown. Cytoplasmic line within each breed of origin was treated as a random effect. Variance components for random effects were estimated using derivative-free REML procedures. Line of cytoplasm accounted for less than .002% of the phenotypic variance in all three traits. Estimates for cytoplasmic breed of origin effects were small in magnitude, and contrasts tested (Angus vs Brahman and Angus vs Unknown) were not significant (P > .10). Estimates of heritability of direct (maternal) effects were .36 (.20), .41 (.27), and .21 (.08) for birth weight, weaning weight, and postweaning average daily gain, respectively. Estimates of genetic correlations between direct and maternal effects ranged from -.27 for postweaning average daily gain to -.58 for birth weight. No evidence for breed or line within breed of cytoplasmic origin effects was detected in these data. PMID:7730171

  16. Indocyanine green as effective antibody conjugate for intracellular molecular targeted photodynamic therapy

    Science.gov (United States)

    Wang, Sijia; Hüttmann, Gereon; Rudnitzki, Florian; Diddens-Tschoeke, Heyke; Zhang, Zhenxi; Rahmanzadeh, Ramtin

    2016-07-01

    The fluorescent dye indocyanine green (ICG) is clinically approved and has been applied for ophthalmic and intraoperative angiography, measurement of cardiac output and liver function, or as contrast agent in cancer surgery. Though ICG is known for its photochemical effects, it has played a minor role so far in photodynamic therapy or techniques for targeted protein-inactivation. Here, we investigated ICG as an antibody-conjugate for the selective inactivation of the protein Ki-67 in the nucleus of cells. Conjugates of the Ki-67 antibody TuBB-9 with different amounts of ICG were synthesized and delivered into HeLa and OVCAR-5 cells through conjugation to the nuclear localization sequence. Endosomal escape of the macromolecular antibodies into the cytoplasm was optically triggered by photochemical internalization with the photosensitizer BPD. The second light irradiation at 690 nm inactivated Ki-67 and subsequently caused cell death. Here, we show that ICG as an antibody-conjugate can be an effective photosensitizing agent. Best effects were achieved with 1.8 ICG molecules per antibody. Conjugated to antibodies, the ICG absorption peaks vary proportionally with concentration. The absorption of ICG above 650 nm within the optical window of tissue opens the possibility of selective Ki-67 inactivation deep inside of tissues.

  17. Indocyanine green as effective antibody conjugate for intracellular molecular targeted photodynamic therapy

    Science.gov (United States)

    Wang, Sijia; Hüttmann, Gereon; Rudnitzki, Florian; Diddens-Tschoeke, Heyke; Zhang, Zhenxi; Rahmanzadeh, Ramtin

    2016-07-01

    The fluorescent dye indocyanine green (ICG) is clinically approved and has been applied for ophthalmic and intraoperative angiography, measurement of cardiac output and liver function, or as contrast agent in cancer surgery. Though ICG is known for its photochemical effects, it has played a minor role so far in photodynamic therapy or techniques for targeted protein-inactivation. Here, we investigated ICG as an antibody-conjugate for the selective inactivation of the protein Ki-67 in the nucleus of cells. Conjugates of the Ki-67 antibody TuBB-9 with different amounts of ICG were synthesized and delivered into HeLa and OVCAR-5 cells through conjugation to the nuclear localization sequence. Endosomal escape of the macromolecular antibodies into the cytoplasm was optically triggered by photochemical internalization with the photosensitizer BPD. The second light irradiation at 690 nm inactivated Ki-67 and subsequently caused cell death. Here, we show that ICG as an antibody-conjugate can be an effective photosensitizing agent. Best effects were achieved with 1.8 ICG molecules per antibody. Conjugated to antibodies, the ICG absorption peaks vary proportionally with concentration. The absorption of ICG above 650 nm within the optical window of tissue opens the possibility of selective Ki-67 inactivation deep inside of tissues.

  18. Discrete nuclear structures in actively growing neuroblastoma cells are revealed by antibodies raised against phosphorylated neurofilament proteins

    Directory of Open Access Journals (Sweden)

    Raabe Timothy D

    2003-04-01

    Full Text Available Abstract Background Nuclear objects that have in common the property of being recognized by monoclonal antibodies specific for phosphoprotein epitopes and cytoplasmic intermediate filaments (in particular, SMI-31 and RT-97 have been reported in glial and neuronal cells, in situ and in vitro. Since neurofilament and glial filaments are generally considered to be restricted to the cytoplasm, we were interested in exploring the identity of the structures labeled in the nucleus as well as the conditions under which they could be found there. Results Using confocal microscopy and western analysis techniques, we determined 1 the immunolabeled structures are truly within the nucleus; 2 the phosphoepitope labeled by SMI-31 and RT-97 is not specific to neurofilaments (NFs and it can be identified on other intermediate filament proteins (IFs in other cell types; and 3 there is a close relationship between DNA synthesis and the amount of nuclear staining by these antibodies thought to be specific for cytoplasmic proteins. Searches of protein data bases for putative phosphorylation motifs revealed that lamins, NF-H, and GFAP each contain a single tyrosine phosphorylation motif with nearly identical amino acid sequence. Conclusion We therefore suggest that this sequence may be the epitope recognized by SMI-31 and RT-97 mABs, and that the nuclear structures previously reported and shown here are likely phosphorylated lamin intermediate filaments, while the cytoplasmic labeling revealed by the same mABs indicates phosphorylated NFs in neurons or GFAP in glia.

  19. Carbimazole-induced agranulocytosis

    Directory of Open Access Journals (Sweden)

    Anisha Mohan

    2015-01-01

    Full Text Available A 47 year old lady with hyperthyroidism for past 1½ years was initially on Carbimazole 20 mg orally then changed to 30 mg (during Hysterectomy but was taking 10 mg for last 1 year. She had intermittent fever with severe B/L bifrontal headache since 3 weeks. Routine investigations showed anaemia, neutropenia, leucopenia and CRP elevation. Peripheral smear showed normocytic normochromic anaemia with Rouleaux formation, leucopenia with 2% atypical cells and mild thrombocytosis. Widal test, RA factor (Rheumatoid factor test, Ig M (Immunoglobulin M dengue, Ig M Lepto, TORCH infections (Toxoplasmosis, Other (Syphilis, varicella-zoster, parvovirus B19, Cytomegalovirus and Herpes infections, ANA (Antinuclear antibody screen cANCA (Cytoplasmic antineutrophil cytoplasmic antibodies and pANCA (Perinuclear Anti-Neutrophil Cytoplasmic Antibodies tests were negative. Bone marrow aspiration showed normo to hypercellular marrow with 15% atypical cells and plasma cells. Multiple myeloma workup was done. Carbimazole was withheld. Conclusion: Drug induced agranulocytosis occurs with in 1-2 months of taking the antithyroid medication but onset delayed by 1½ year. De-challenge resulted normalization of blood parameters.

  20. Probing Functional Changes in Exocyst Configuration with Monoclonal Antibodies.

    Science.gov (United States)

    Inamdar, Shivangi M; Hsu, Shu-Chan; Yeaman, Charles

    2016-01-01

    . Antibodies to Sec6NT co-precipitated substantially more Sec5, -10, -15, Exo70 and -84 than did those to Sec6CT. In contrast, antibodies to Sec6CT co-precipitated more Sec3 and Sec8 than did those to Sec6NT. These results are consistent with a model in which exocyst activation during periods of rapid membrane expansion is accompanied by molecular rearrangements within the holocomplex or association with accessory proteins, which expose the Sec6 C-terminal domain when the complex is membrane-bound and conceal it when the complex is cytoplasmic. PMID:27376061

  1. Loss of Tropomodulin4 in the zebrafish mutant träge causes cytoplasmic rod formation and muscle weakness reminiscent of nemaline myopathy

    Directory of Open Access Journals (Sweden)

    Joachim Berger

    2014-12-01

    Full Text Available Nemaline myopathy is an inherited muscle disease that is mainly diagnosed by the presence of nemaline rods in muscle biopsies. Of the nine genes associated with the disease, five encode components of striated muscle sarcomeres. In a genetic zebrafish screen, the mutant träge (trg was isolated based on its reduction in muscle birefringence, indicating muscle damage. Myofibres in trg appeared disorganised and showed inhomogeneous cytoplasmic eosin staining alongside malformed nuclei. Linkage analysis of trg combined with sequencing identified a nonsense mutation in tropomodulin4 (tmod4, a regulator of thin filament length and stability. Accordingly, although actin monomers polymerize to form thin filaments in the skeletal muscle of tmod4trg mutants, thin filaments often appeared to be dispersed throughout myofibres. Organised myofibrils with the typical striation rarely assemble, leading to severe muscle weakness, impaired locomotion and early death. Myofibrils of tmod4trg mutants often featured thin filaments of various lengths, widened Z-disks, undefined H-zones and electron-dense aggregations of various shapes and sizes. Importantly, Gomori trichrome staining and the lattice pattern of the detected cytoplasmic rods, together with the reactivity of rods with phalloidin and an antibody against actinin, is reminiscent of nemaline rods found in nemaline myopathy, suggesting that misregulation of thin filament length causes cytoplasmic rod formation in tmod4trg mutants. Although Tropomodulin4 has not been associated with myopathy, the results presented here implicateTMOD4 as a novel candidate for unresolved nemaline myopathies and suggest that the tmod4trg mutant will be a valuable tool to study human muscle disorders.

  2. Thymosin Beta 4 May Translocate from the Cytoplasm in to the Nucleus in HepG2 Cells following Serum Starvation. An Ultrastructural Study

    Science.gov (United States)

    Piludu, Marco; Piras, Monica; Pichiri, Giuseppina; Coni, Pierpaolo; Orrù, Germano; Cabras, Tiziana; Messana, Irene; Faa, Gavino; Castagnola, Massimo

    2015-01-01

    Due to its actin-sequestering properties, thymosin beta-4 (Tβ4) is considered to play a significant role in the cellular metabolism. Several physiological properties of Tβ4 have been reported;, however, many questions concerning its cellular function remain to be ascertained. To better understand the role of this small peptide we have analyzed by means of transmission immunoelectron microscopy techniques the ultrastructural localization of Tβ4 in HepG2 cells. Samples of HepG2 cells were fixed in a mixture of 3% formaldehyde and 0.1% glutaraldehyde in 0.1 M cacodylate buffer and processed for standard electron microscopic techniques. The samples were dehydrated in a cold graded methanol series and embedded in LR gold resin. Ultrathin sections were labeled with rabbit antibodies to Tβ4, followed by gold-labeled goat anti-rabbit, stained with uranyl acetate and bismuth subnitrate, observed and photographed in a JEOL 100S transmission electron microscope. High-resolution electron microscopy showed that Tβ4 was mainly restricted to the cytoplasm of HepG2 growing in complete medium. A strong Tβ4 reactivity was detected in the perinuclear region of the cytoplasmic compartment where gold particles appeared strictly associated to the nuclear membrane. In the nucleus specific Tβ4 labeling was observed in the nucleolus. The above electron microscopic results confirm and extend previous observations at light microscopic level, highlighting the subcellular distribution of Tβ4 in both cytoplasmic and nuclear compartments of HepG2 cells. The meaning of Tβ4 presence in the nucleolus is not on the best of our knowledge clarified yet. It could account for the interaction of Tβ4 with nucleolar actin and according with this hypothesis, Tβ4 could contribute together with the other nucleolar acting binding proteins to modulate the transcription activity of the RNA polymerases. PMID:25835495

  3. Cytoplasmic p21 is a potential predictor for cisplatin sensitivity in ovarian cancer

    Directory of Open Access Journals (Sweden)

    Liu Ronghua

    2011-09-01

    Full Text Available Abstract Background P21(WAF1/Cip1 binds to cyclin-dependent kinase complexes and inhibits their activities. It was originally described as an inhibitor of cancer cell proliferation. However, many recent studies have shown that p21 promotes tumor progression when accumulated in the cell cytoplasm. So far, little is known about the correlation between cytoplasmic p21 and drug resistance. This study was aimed to investigate the role of p21 in the cisplatin resistance of ovarian cancer. Methods RT-PCR, western blot and immunofluorescence were used to detect p21 expression and location in cisplatin-resistant ovarian cancer cell line C13* and its parental line OV2008. Regulation of cytoplasmic p21 was performed through transfection of p21 siRNA, Akt2 shRNA and Akt2 constitutively active vector in the two cell lines; their effects on cisplatin-induced apoptosis were evaluated by flow cytometry. Tumor tissue sections of clinical samples were analyzed by immunohistochemistry. Results p21 predominantly localizes to the cytoplasm in C13* compared to OV2008. Persistent exposure to low dose cisplatin in OV2008 leads to p21 translocation from nuclear to cytoplasm, while it had not impact on p21 localization in C13*. Knockdown of cytoplasmic p21 by p21 siRNA transfection in C13* notably increased cisplatin-induced apoptosis through activation of caspase 3. Inhibition of p21 translocation into the cytoplasm by transfection of Akt2 shRNA into C13* cells significantly increased cisplatin-induced apoptosis, while induction of p21 translocation into the cytoplasm by transfection of constitutively active Akt2 in OV2008 enhanced the resistance to cisplatin. Immunohistochemical analysis of clinical ovarian tumor tissues demonstrated that cytoplasmic p21 was negatively correlated with the response to cisplatin based treatment. Conclusions Cytoplasmic p21 is a novel biomarker of cisplatin resistance and it may represent a potential therapeutic target for ovarian tumors

  4. Targeting of Antibodies using Aptamers

    OpenAIRE

    Missailidis, Sotiris

    2003-01-01

    The chapter presents a methodology for the rapid selection of aptamers against antibody targets. It is a detailed account of the various methodological steps that describe the selection of aptamers, including PCR steps, buffers to be used, target immobilisation, partitioning and amplification of aptamers, clonning and sequencing, to results in high affinity and specificity ligands for the chosen target antibody.

  5. Structural Characterization of Peptide Antibodies

    DEFF Research Database (Denmark)

    Chailyan, Anna; Marcatili, Paolo

    2015-01-01

    The role of proteins as very effective immunogens for the generation of antibodies is indisputable. Nevertheless, cases in which protein usage for antibody production is not feasible or convenient compelled the creation of a powerful alternative consisting of synthetic peptides. Synthetic peptide...

  6. Pathogenic role of antiphospholipid antibodies

    NARCIS (Netherlands)

    Salmon, J. E.; de Groot, P. G.

    2008-01-01

    The antiphospholipid antibody syndrome (APS) is characterized by recurrent arterial and venous thrombosis and/or pregnancy in association with antiphospholipid (aPL) antibodies. The pathogenic mechanisms in APS that lead to in vivo injury are incompletely understood. Recent evidence suggests that AP

  7. Metrics for antibody therapeutics development.

    Science.gov (United States)

    Reichert, Janice M

    2010-01-01

    A wide variety of full-size monoclonal antibodies (mAbs) and therapeutics derived from alternative antibody formats can be produced through genetic and biological engineering techniques. These molecules are now filling the preclinical and clinical pipelines of every major pharmaceutical company and many biotechnology firms. Metrics for the development of antibody therapeutics, including averages for the number of candidates entering clinical study and development phase lengths for mAbs approved in the United States, were derived from analysis of a dataset of over 600 therapeutic mAbs that entered clinical study sponsored, at least in part, by commercial firms. The results presented provide an overview of the field and context for the evaluation of on-going and prospective mAb development programs. The expansion of therapeutic antibody use through supplemental marketing approvals and the increase in the study of therapeutics derived from alternative antibody formats are discussed. PMID:20930555

  8. Metrics for antibody therapeutics development.

    Science.gov (United States)

    Reichert, Janice M

    2010-01-01

    A wide variety of full-size monoclonal antibodies (mAbs) and therapeutics derived from alternative antibody formats can be produced through genetic and biological engineering techniques. These molecules are now filling the preclinical and clinical pipelines of every major pharmaceutical company and many biotechnology firms. Metrics for the development of antibody therapeutics, including averages for the number of candidates entering clinical study and development phase lengths for mAbs approved in the United States, were derived from analysis of a dataset of over 600 therapeutic mAbs that entered clinical study sponsored, at least in part, by commercial firms. The results presented provide an overview of the field and context for the evaluation of on-going and prospective mAb development programs. The expansion of therapeutic antibody use through supplemental marketing approvals and the increase in the study of therapeutics derived from alternative antibody formats are discussed.

  9. Bayesian Inference of Forces Causing Cytoplasmic Streaming in Caenorhabditis elegans Embryos and Mouse Oocytes.

    Science.gov (United States)

    Niwayama, Ritsuya; Nagao, Hiromichi; Kitajima, Tomoya S; Hufnagel, Lars; Shinohara, Kyosuke; Higuchi, Tomoyuki; Ishikawa, Takuji; Kimura, Akatsuki

    2016-01-01

    Cellular structures are hydrodynamically interconnected, such that force generation in one location can move distal structures. One example of this phenomenon is cytoplasmic streaming, whereby active forces at the cell cortex induce streaming of the entire cytoplasm. However, it is not known how the spatial distribution and magnitude of these forces move distant objects within the cell. To address this issue, we developed a computational method that used cytoplasm hydrodynamics to infer the spatial distribution of shear stress at the cell cortex induced by active force generators from experimentally obtained flow field of cytoplasmic streaming. By applying this method, we determined the shear-stress distribution that quantitatively reproduces in vivo flow fields in Caenorhabditis elegans embryos and mouse oocytes during meiosis II. Shear stress in mouse oocytes were predicted to localize to a narrower cortical region than that with a high cortical flow velocity and corresponded with the localization of the cortical actin cap. The predicted patterns of pressure gradient in both species were consistent with species-specific cytoplasmic streaming functions. The shear-stress distribution inferred by our method can contribute to the characterization of active force generation driving biological streaming.

  10. Bayesian Inference of Forces Causing Cytoplasmic Streaming in Caenorhabditis elegans Embryos and Mouse Oocytes

    Science.gov (United States)

    Niwayama, Ritsuya; Nagao, Hiromichi; Kitajima, Tomoya S.; Hufnagel, Lars; Shinohara, Kyosuke; Higuchi, Tomoyuki; Ishikawa, Takuji

    2016-01-01

    Cellular structures are hydrodynamically interconnected, such that force generation in one location can move distal structures. One example of this phenomenon is cytoplasmic streaming, whereby active forces at the cell cortex induce streaming of the entire cytoplasm. However, it is not known how the spatial distribution and magnitude of these forces move distant objects within the cell. To address this issue, we developed a computational method that used cytoplasm hydrodynamics to infer the spatial distribution of shear stress at the cell cortex induced by active force generators from experimentally obtained flow field of cytoplasmic streaming. By applying this method, we determined the shear-stress distribution that quantitatively reproduces in vivo flow fields in Caenorhabditis elegans embryos and mouse oocytes during meiosis II. Shear stress in mouse oocytes were predicted to localize to a narrower cortical region than that with a high cortical flow velocity and corresponded with the localization of the cortical actin cap. The predicted patterns of pressure gradient in both species were consistent with species-specific cytoplasmic streaming functions. The shear-stress distribution inferred by our method can contribute to the characterization of active force generation driving biological streaming. PMID:27472658

  11. Cytoplasmic calcium levels in protoplasts from the cap and elongation zone of maize roots

    Science.gov (United States)

    Kiss, H. G.; Evans, M. L.; Johnson, J. D.

    1991-01-01

    Calcium has been implicated as a key component in the signal transduction process of root gravitropism. We measured cytoplasmic free calcium in protoplasts isolated from the elongation zone and cap of primary roots of light-grown, vertically oriented seedlings of Zea mays L. Protoplasts were loaded with the penta-potassium salts of fura-2 and indo-1 by incubation in acidic solutions of these calcium indicators. Loading increased with decreasing pH but the pH dependence was stronger for indo-1 than for fura-2. In the case of fura-2, loading was enhanced only at the lowest pH (4.5) tested. Dyes loaded in this manner were distributed predominantly in the cytoplasm as indicated by fluorescence patterns. As an alternative method of loading, protoplasts were incubated with the acetoxymethylesters of fura-2 and indo-1. Protoplasts loaded by this method exhibited fluorescence both in the cytoplasm and in association with various organelles. Cytoplasmic calcium levels measured using spectrofluorometry, were found to be 160 +/- 40 nM and 257 +/- 27 nM, respectively, in populations of protoplasts from the root cap and elongation zone. Cytoplasmic free calcium did not increase upon addition of calcium to the incubation medium, indicating that the passive permeability to calcium was low.

  12. Regulation of Actin-Dependent Cytoplasmic Motility by Type II Phytochrome Occurs within Seconds in Vallisneria gigantea Epidermal Cells

    Science.gov (United States)

    Takagi, Shingo; Kong, Sam-Geun; Mineyuki, Yoshinobu; Furuya, Masaki

    2003-01-01

    The effects of light on actin-dependent cytoplasmic motility in epidermal cells of green leaves of the aquatic angiosperm Vallisneria gigantea were investigated quantitatively using a custom-made dynamic image analyzer. Cytoplasmic motility was measured by monitoring changes in the brightness of individual pixels on digitized images taken sequentially under infrared light. Acceleration and deceleration of cytoplasmic motility were regulated photoreversibly by type II phytochrome(s). This phytochrome-dependent induction of cytoplasmic motility did not occur uniformly in cytoplasm but took place as scattered patches in which no particular organelles, including nucleus, existed. The induction became detectable at 2.5 s after the start of irradiation with pulsed red light. In cells exposed to microbeam irradiation, cytoplasmic motility was induced only in sites in the cytoplasm that were irradiated directly, whereas nonirradiated neighboring areas were unaffected. The effect was short-lived, disappearing within a few minutes, and no signal was transmitted from an irradiated cell to its neighbors. Anti-phytochrome antibody–responsive protein(s) was detectable in the leaf extract by immunoblot and zinc blot analyses and in cryosections of the epidermis by immunocytochemistry. Although the phytochrome-dependent cytoplasmic motility was blocked by exogenously applied latrunculin B or cytochalasins, treatment of the dark-adapted cells with Ca2+-chelating reagents induced the cytoplasmic motility. We have proposed a model for the phytochrome regulation of cytoplasmic motility as one of the earliest responses to a light stimulus. PMID:12566576

  13. Cytoplasmic Skp2 expression is associated with p-Akt1 and predicts poor prognosis in human breast carcinomas.

    Directory of Open Access Journals (Sweden)

    Jing Liu

    Full Text Available BACKGROUND: S-phase kinase protein 2 (Skp2, an oncogenic protein, is a key regulator in different cellular and molecular processes, through ubiquitin-proteasome degradation pathway. Increased levels of Skp2 are observed in various types of cancer and associated with poor prognosis. However, in human breast carcinomas, the underlying mechanism and prognostic significance of cytoplasmic Skp2 is still undefined. METHODS: To investigate the role of cytoplasmic Skp2 expression in human breast carcinomas, we immnohistochemically assessed cytoplasmic Skp2, p-Akt1, and p27 expression in 251 patients with invasive ductal carcinomas of the breast. Association of cytoplasmic Skp2 expression with p-Akt1 and p27 was analyzed as well as correspondence with other clinicopathological parameters. Disease-free survival and overall survival were determined based on the Kaplan-Meier method and Cox regression models. RESULTS: Cytoplasmic of Skp2 was detected in 165 out of 251 (65.7% patients. Cytoplasmic Skp2 expression was associated with larger tumor size, more advanced histological grade, and positive HER2 expression. Increased cytoplasmic Skp2 expression correlated with p-Akt1 expression, with 54.2% (51/94 of low p-Akt1-expressing breast carcinomas, but 72.6% (114/157 of high p-Akt1-expressing breast carcinomas exhibiting cytoplasmic Skp2 expression. Elevated cytoplasmic Skp2 expression with low p-Akt1 expression was associated with poor disease-free and overall survival (DFS and OS, and Cox regression models demonstrated that cytoplasmic Skp2 expression was an independent prognostic marker for invasive breast carcinomas. CONCLUSION: Cytoplasmic Skp2 expression is associated with aggressive prognostic factors, such as larger tumor size, and advanced histological grade of the breast cancers. Results demonstrate that combined cytoplasmic Skp2 and p-Akt1 expression may be prognostic for patients with invasive breast carcinomas, and cytoplasmic Skp2 may serve as a

  14. An FNA pitfall: Mammary analog secretory carcinoma mistaken for acinic cell carcinoma due to cytoplasmic granules

    Directory of Open Access Journals (Sweden)

    Nouf Hijazi, MD

    2014-12-01

    Full Text Available In the salivary gland, a key differential feature of Mammary analog secretory carcinoma (MASC from acinic cell carcinoma (ACC is the lack of cytoplasmic granules. We report a case of a parotid mass incorrectly diagnosed on fine needle aspirate as acinic cell carcinoma due to many cells with basophilic granules suggesting serous acinar differention. Tumor resection revealed a tumor consistent with low grade adenocarcinoma that had eosinophilic, microvacuolar cytoplasm with distinct basophilic granules staining with PASD and mucicarmine. The diagnosis of MASC was confirmed with stains for GCDF-15, mammoglobin, and S100 and FISH consistent with a t(12;15 translocation. Relying on the absence of cytoplasmic granules as a feature to distinguish ACC from MASC is a diagnostic pitfall.

  15. Vesicular Nucleo-Cytoplasmic Transport—Herpesviruses as Pioneers in Cell Biology

    Science.gov (United States)

    Mettenleiter, Thomas C.

    2016-01-01

    Herpesviruses use a vesicle-mediated transfer of intranuclearly assembled nucleocapsids through the nuclear envelope (NE) for final maturation in the cytoplasm. The molecular basis for this novel vesicular nucleo-cytoplasmic transport is beginning to be elucidated in detail. The heterodimeric viral nuclear egress complex (NEC), conserved within the classical herpesviruses, mediates vesicle formation from the inner nuclear membrane (INM) by polymerization into a hexagonal lattice followed by fusion of the vesicle membrane with the outer nuclear membrane (ONM). Mechanisms of capsid inclusion as well as vesicle-membrane fusion, however, are largely unclear. Interestingly, a similar transport mechanism through the NE has been demonstrated in nuclear export of large ribonucleoprotein complexes during Drosophila neuromuscular junction formation, indicating a widespread presence of a novel concept of cellular nucleo-cytoplasmic transport. PMID:27690080

  16. Dynamics of highly polydisperse colloidal suspensions as a model system for bacterial cytoplasm

    Science.gov (United States)

    Hwang, Jiye; Kim, Jeongmin; Sung, Bong June

    2016-08-01

    There are various kinds of macromolecules in bacterial cell cytoplasm. The size polydispersity of the macromolecules is so significant that the crystallization and the phase separation could be suppressed, thus stabilizing the liquid state of bacterial cytoplasm. On the other hand, recent experiments suggested that the macromolecules in bacterial cytoplasm should exhibit glassy dynamics, which should be also affected significantly by the size polydispersity of the macromolecules. In this work, we investigate the anomalous and slow dynamics of highly polydisperse colloidal suspensions, of which size distribution is chosen to mimic Escherichia coli cytoplasm. We find from our Langevin dynamics simulations that the diffusion coefficient (Dtot) and the displacement distribution functions (P (r ,t ) ) averaged over all colloids of different sizes do not show anomalous and glassy dynamic behaviors until the system volume fraction ϕ is increased up to 0.82. This indicates that the intrinsic polydispersity of bacterial cytoplasm should suppress the glass transition and help maintain the liquid state of the cytoplasm. On the other hand, colloids of each kind show totally different dynamic behaviors depending on their size. The dynamics of colloids of different size becomes non-Gaussian at a different range of ϕ , which suggests that a multistep glass transition should occur. The largest colloids undergo the glass transition at ϕ =0.65 , while the glass transition does not occur for smaller colloids in our simulations even at the highest value of ϕ . We also investigate the distribution (P (θ ,t ) ) of the relative angles of displacement for macromolecules and find that macromolecules undergo directionally correlated motions in a sufficiently dense system.

  17. Distinct cytoplasmic and nuclear functions of the stress induced protein DDIT3/CHOP/GADD153.

    Directory of Open Access Journals (Sweden)

    Alexandra Jauhiainen

    Full Text Available DDIT3, also known as GADD153 or CHOP, encodes a basic leucine zipper transcription factor of the dimer forming C/EBP family. DDIT3 is known as a key regulator of cellular stress response, but its target genes and functions are not well characterized. Here, we applied a genome wide microarray based expression analysis to identify DDIT3 target genes and functions. By analyzing cells carrying tamoxifen inducible DDIT3 expression constructs we show distinct gene expression profiles for cells with cytoplasmic and nuclear localized DDIT3. Of 175 target genes identified only 3 were regulated by DDIT3 in both cellular localizations. More than two thirds of the genes were downregulated, supporting a role for DDIT3 as a dominant negative factor that could act by either cytoplasmic or nuclear sequestration of dimer forming transcription factor partners. Functional annotation of target genes showed cell migration, proliferation and apoptosis/survival as the most affected categories. Cytoplasmic DDIT3 affected more migration associated genes, while nuclear DDIT3 regulated more cell cycle controlling genes. Cell culture experiments confirmed that cytoplasmic DDIT3 inhibited migration, while nuclear DDIT3 caused a G1 cell cycle arrest. Promoters of target genes showed no common sequence motifs, reflecting that DDIT3 forms heterodimers with several alternative transcription factors that bind to different motifs. We conclude that expression of cytoplasmic DDIT3 regulated 94 genes. Nuclear translocation of DDIT3 regulated 81 additional genes linked to functions already affected by cytoplasmic DDIT3. Characterization of DDIT3 regulated functions helps understanding its role in stress response and involvement in cancer and degenerative disorders.

  18. Day-to-night variations of cytoplasmic pH in a crassulacean acid metabolism plant.

    Science.gov (United States)

    Hafke, J B; Neff, R; Hütt, M T; Lüttge, U; Thiel, G

    2001-01-01

    In crassulacean acid metabolism (CAM) large amounts of malic acid are redistributed between vacuole and cytoplasm in the course of night-to-day transitions. The corresponding changes of the cytoplasmic pH (pHcyt) were monitored in mesophyll protoplasts from the CAM plant Kalanchoe daigremontiana Hamet et Perrier by ratiometric fluorimetry with the fluorescent dye 2',7'-bis-(2-carboxyethyl)-5-(and-6-)carboxyfluorescein as a pHcyt indicator. At the beginning of the light phase, pHcyt was slightly alkaline (about 7.5). It dropped during midday by about 0.3 pH units before recovering again in the late-day-to-early-dark phase. In the physiological context the variation in pHcyt may be a component of CAM regulation. Due to its pH sensitivity, phosphoenolpyruvate carboxylase appears as a likely target enzyme. From monitoring delta pHcyt in response to loading the cytoplasm with the weak acid salt K-acetate a cytoplasmic H(+)-buffer capacity in the order of 65 mM H+ per pH unit was estimated at a pHcyt of about 7.5. With this value, an acid load of the cytoplasm by about 10 mM malic acid can be estimated as the cause of the observed drop in pHcyt. A diurnal oscillation in pHcyt and a quantitatively similar cytoplasmic malic acid is predicted from an established mathematical model which allows simulation of the CAM dynamics. The similarity of model predictions and experimental data supports the view put forward in this model that a phase transition of the tonoplast is an essential functional element in CAM dynamics. PMID:11732184

  19. A Mechanism for Cytoplasmic Streaming: Kinesin-Driven Alignment of Microtubules and Fast Fluid Flows.

    Science.gov (United States)

    Monteith, Corey E; Brunner, Matthew E; Djagaeva, Inna; Bielecki, Anthony M; Deutsch, Joshua M; Saxton, William M

    2016-05-10

    The transport of cytoplasmic components can be profoundly affected by hydrodynamics. Cytoplasmic streaming in Drosophila oocytes offers a striking example. Forces on fluid from kinesin-1 are initially directed by a disordered meshwork of microtubules, generating minor slow cytoplasmic flows. Subsequently, to mix incoming nurse cell cytoplasm with ooplasm, a subcortical layer of microtubules forms parallel arrays that support long-range, fast flows. To analyze the streaming mechanism, we combined observations of microtubule and organelle motions with detailed mathematical modeling. In the fast state, microtubules tethered to the cortex form a thin subcortical layer and undergo correlated sinusoidal bending. Organelles moving in flows along the arrays show velocities that are slow near the cortex and fast on the inward side of the subcortical microtubule layer. Starting with fundamental physical principles suggested by qualitative hypotheses, and with published values for microtubule stiffness, kinesin velocity, and cytoplasmic viscosity, we developed a quantitative coupled hydrodynamic model for streaming. The fully detailed mathematical model and its simulations identify key variables that can shift the system between disordered (slow) and ordered (fast) states. Measurements of array curvature, wave period, and the effects of diminished kinesin velocity on flow rates, as well as prior observations on f-actin perturbation, support the model. This establishes a concrete mechanistic framework for the ooplasmic streaming process. The self-organizing fast phase is a result of viscous drag on kinesin-driven cargoes that mediates equal and opposite forces on cytoplasmic fluid and on microtubules whose minus ends are tethered to the cortex. Fluid moves toward plus ends and microtubules are forced backward toward their minus ends, resulting in buckling. Under certain conditions, the buckling microtubules self-organize into parallel bending arrays, guiding varying directions

  20. Analysis of cytoplasmic effects and fine-mapping of a genic male sterile line in rice.

    Directory of Open Access Journals (Sweden)

    Peng Qin

    Full Text Available Cytoplasm has substantial genetic effects on progeny and is important for yield improvement in rice breeding. Studies on the cytoplasmic effects of cytoplasmic male sterility (CMS show that most types of CMS have negative effects on yield-related traits and that these negative effects vary among CMS. Some types of genic male sterility (GMS, including photo-thermo sensitive male sterility (PTMS, have been widely used in rice breeding, but the cytoplasmic effects of GMS remain unknown. Here, we identified a GMS mutant line, h2s, which exhibited small, white anthers and failed to produce mature pollen. Unlike CMS, the h2s had significant positive cytoplasmic effects on the seed set rate, weight per panicle, yield, and general combining ability (GCA for plant height, seed set rate, weight per panicle, and yield. These effects indicated that h2s cytoplasm may show promise for the improvement of rice yield. Genetic analysis suggested that the phenotype of h2s was controlled by a single recessive locus. We mapped h2s to a 152 kb region on chromosome 6, where 22 candidate genes were predicted. None of the 22 genes had previously been reported to be responsible for the phenotypes of h2s. Sequencing analysis showed a 12 bp deletion in the sixth exon of Loc_Os06g40550 in h2s in comparison to wild type, suggesting that Loc_Os06g40550 is the best candidate gene. These results lay a strong foundation for cloning of the H2S gene to elucidate the molecular mechanism of male reproduction.

  1. Transcriptomic Changes Due to Cytoplasmic TDP-43 Expression Reveal Dysregulation of Histone Transcripts and Nuclear Chromatin.

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    Alexandre Amlie-Wolf

    Full Text Available TAR DNA-binding protein 43 (TDP-43 is normally a nuclear RNA-binding protein that exhibits a range of functions including regulation of alternative splicing, RNA trafficking, and RNA stability. However, in amyotrophic lateral sclerosis (ALS and frontotemporal lobar degeneration with TDP-43 inclusions (FTLD-TDP, TDP-43 is abnormally phosphorylated, ubiquitinated, and cleaved, and is mislocalized to the cytoplasm where it forms distinctive aggregates. We previously developed a mouse model expressing human TDP-43 with a mutation in its nuclear localization signal (ΔNLS-hTDP-43 so that the protein preferentially localizes to the cytoplasm. These mice did not exhibit a significant number of cytoplasmic aggregates, but did display dramatic changes in gene expression as measured by microarray, suggesting that cytoplasmic TDP-43 may be associated with a toxic gain-of-function. Here, we analyze new RNA-sequencing data from the ΔNLS-hTDP-43 mouse model, together with published RNA-sequencing data obtained previously from TDP-43 antisense oligonucleotide (ASO knockdown mice to investigate further the dysregulation of gene expression in the ΔNLS model. This analysis reveals that the transcriptomic effects of the overexpression of the ΔNLS-hTDP-43 transgene are likely due to a gain of cytoplasmic function. Moreover, cytoplasmic TDP-43 expression alters transcripts that regulate chromatin assembly, the nucleolus, lysosomal function, and histone 3' untranslated region (UTR processing. These transcriptomic alterations correlate with observed histologic abnormalities in heterochromatin structure and nuclear size in transgenic mouse and human brains.

  2. Comparative assessment of genetic diversity in cytoplasmic and nuclear genome of upland cotton.

    Science.gov (United States)

    Egamberdiev, Sharof S; Saha, Sukumar; Salakhutdinov, Ilkhom; Jenkins, Johnie N; Deng, Dewayne; Y Abdurakhmonov, Ibrokhim

    2016-06-01

    The importance of the cytoplasmic genome for many economically important traits is well documented in several crop species, including cotton. There is no report on application of cotton chloroplast specific SSR markers as a diagnostic tool to study genetic diversity among improved Upland cotton lines. The complete plastome sequence information in GenBank provided us an opportunity to report on 17 chloroplast specific SSR markers using a cost-effective data mining strategy. Here we report the comparative analysis of genetic diversity among a set of 42 improved Upland cotton lines using SSR markers specific to chloroplast and nuclear genome, respectively. Our results revealed that low to moderate level of genetic diversity existed in both nuclear and cytoplasm genome among this set of cotton lines. However, the specific estimation suggested that genetic diversity is lower in cytoplasmic genome compared to the nuclear genome among this set of Upland cotton lines. In summary, this research is important from several perspectives. We detected a set of cytoplasm genome specific SSR primer pairs by using a cost-effective data mining strategy. We reported for the first time the genetic diversity in the cytoplasmic genome within a set of improved Upland cotton accessions. Results revealed that the genetic diversity in cytoplasmic genome is narrow, compared to the nuclear genome within this set of Upland cotton accessions. Our results suggested that most of these polymorphic chloroplast SSRs would be a valuable complementary tool in addition to the nuclear SSR in the study of evolution, gene flow and genetic diversity in Upland cotton.

  3. Laboratory tests for the diagnosis and management of chronic canine and feline enteropathies.

    Science.gov (United States)

    Berghoff, Nora; Steiner, Jörg M

    2011-03-01

    Chronic enteropathies are commonly encountered in both cats and dogs. Although definitive diagnosis often requires collection of gastrointestinal biopsies for histopathologic evaluation, less invasive laboratory tests can be highly informative and should be performed prior to biopsy collection. Tests for determination of infectious causes comprise those for helminthic, protozoal, bacterial, or fungal organisms. Intestinal function and disease may be assessed by measuring serum concentrations of cobalamin, folate, and C-reactive protein, and fecal concentrations of α(1)-proteinase inhibitor. Ongoing research has led to development of tests for serum perinuclear antineutrophilic cytoplasmic antibodies, and fecal inflammatory markers, including S100-proteins and N-methylhistamine.

  4. Meningeal involvement in Behcet's disease: MRI

    International Nuclear Information System (INIS)

    Behcet's disease is a multisystem disease that involves the central nervous system up to half of cases. Presentation with neurologic symptoms occurs in 5 % of cases and cerebral venous thrombosis is one of its major manifestations. A feature not previously reported is progressive meningeal thickening with involvement of both optic nerves. We report a patient with cerebral venous thrombosis, meningeal thickening and contrast enhancement on MRI. This patient had two other unusual features: positive antineutrophil cytoplasmic antibodies and later development of central diabetes insipidus. (orig.)

  5. Coexistence of granulomatosis with polyangiitis (GPA) and Crohn's disease or multiorgan manifestation of the same disease?

    Science.gov (United States)

    Jóźwiak, Lucyna; Ławnicka, Izabela; Książek, Andrzej

    2016-01-01

    Granulomatosis with polyangiitis (GPA) is a systemic necrotizing vasculitis of unknown aetiology, often related to the antineutrophil cytoplasmic antibody (ANCA). GPA was previously named Wegener's granulomatosis (WG). The disease frequently has multisystemic presentation, targeting mainly the respiratory tract and kidneys, but gastrointestinal involvement is uncommon. Crohn's disease (CD) is an inflammatory bowel disease (IBD) with many extraintestinal manifestations. Clinically, symptoms of WG and CD can mimic each other. In this paper a case of GPA manifested initially by severe multiorgan damage including colitis, regarded to be coexistent CD, is presented. The case illustrates the difficulties in establishing the diagnosis when symptoms of the diseases mimic each other.

  6. Monoclonal anti-vasopressin (VP) antibodies penetrate into VP neurons, in vivo.

    Science.gov (United States)

    Burlet, A J; Leon-Henri, B P; Robert, F R; Arahmani, A; Fernette, B M; Burlet, C R

    1987-01-01

    The fate of monoclonal anti-vasopressin antibodies (VP-MAbs) injected in vivo into the paraventricular nucleus (PVN) of the rat brain was studied by immunocytochemistry. Depending on the post survival time, VP-MAbs contained in an ascites fluid were stained at different levels of the VP neurons: the cytoplasm of the PVN neurons, the fibres of the median eminence and the granular layer of the Gyrus Dentatus. The identification of endogenous peptides synthesized by PVN neurons showed that the VP-MAbs uptake was specific: it did not appear either in the oxytocinergic neurons or in the non immunoreactive neurons of the Brattleboro rat brain, this rat being genetically incapable of synthesizing central VP. Conversely, VP-MAbs only penetrated into the VP neurons: ascites fluid containing monoclonal antibodies prepared against bovine thyroglobulin (the carrier conjugated to VP in our immunizations) was neither stained in magnocellular neurons nor carried in nerve fibres. The neuronal uptake and transport of VP-MAbs occurred in vivo: they were totally inhibited by heating of the ascites fluid at 56 degrees C for 30 min; this treatment did not alter the VP-MAbs themselves but probably destroyed some thermic sensitive component essential to the macromolecule internalization. The biological effects of antibodies injected in vivo have been reported. The results described here suggest that some specific antibodies passively transferred into the brain could act directly on the peptide synthesis recognized by the antibodies. PMID:3556490

  7. Conservation and divergence between cytoplasmic and muscle-specific actin capping proteins: insights from the crystal structure of cytoplasmic Cap32/34 from Dictyostelium discoideum

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    Eckert Christian

    2012-06-01

    Full Text Available Abstract Background Capping protein (CP, also known as CapZ in muscle cells and Cap32/34 in Dictyostelium discoideum, plays a major role in regulating actin filament dynamics. CP is a ubiquitously expressed heterodimer comprising an α- and β-subunit. It tightly binds to the fast growing end of actin filaments, thereby functioning as a “cap” by blocking the addition and loss of actin subunits. Vertebrates contain two somatic variants of CP, one being primarily found at the cell periphery of non-muscle tissues while the other is mainly localized at the Z-discs of skeletal muscles. Results To elucidate structural and functional differences between cytoplasmic and sarcomercic CP variants, we have solved the atomic structure of Cap32/34 (32 = β- and 34 = α-subunit from the cellular slime mold Dictyostelium at 2.2 Å resolution and compared it to that of chicken muscle CapZ. The two homologs display a similar overall arrangement including the attached α-subunit C-terminus (α-tentacle and the flexible β-tentacle. Nevertheless, the structures exhibit marked differences suggesting considerable structural flexibility within the α-subunit. In the α-subunit we observed a bending motion of the β-sheet region located opposite to the position of the C-terminal β-tentacle towards the antiparallel helices that interconnect the heterodimer. Recently, a two domain twisting attributed mainly to the β-subunit has been reported. At the hinge of these two domains Cap32/34 contains an elongated and highly flexible loop, which has been reported to be important for the interaction of cytoplasmic CP with actin and might contribute to the more dynamic actin-binding of cytoplasmic compared to sarcomeric CP (CapZ. Conclusions The structure of Cap32/34 from Dictyostelium discoideum allowed a detailed analysis and comparison between the cytoplasmic and sarcomeric variants of CP. Significant structural flexibility could particularly be found within the

  8. Differential localization of cytoplasmic myosin II isoforms A and B in avian interphase and dividing embryonic and immortalized cardiomyocytes and other cell types in vitro

    Science.gov (United States)

    Conrad, A. H.; Jaffredo, T.; Conrad, G. W.; Spooner, B. S. (Principal Investigator)

    1995-01-01

    Two principal isoforms of cytoplasmic myosin II, A and B (CMIIA and CMIIB), are present in different proportions in different tissues. Isoform-specific monoclonal and polyclonal antibodies to avian CMIIA and CMIIB reveal the cellular distributions of these isoforms in interphase and dividing embryonic avian cardiac, intestinal epithelial, spleen, and dorsal root ganglia cells in primary cell culture. Embryonic cardiomyocytes react with antibodies to CMIIB but not to CMIIA, localize CMIIB in stress-fiber-like-structures during interphase, and markedly concentrate CMIIB in networks in the cleavage furrow during cytokinesis. In contrast, cardiac fibroblasts localize both CMIIA and CMIIB in stress fibers and networks during interphase, and demonstrate slight and independently regulated concentration of CMIIA and CMIIB in networks in their cleavage furrows. V-myc-immortalized cardiomyocytes, an established cell line, have regained the ability to express CMIIA, as well as CMIIB, and localize both CMIIA and CMIIB in stress fibers and networks in interphase cells and in cleavage furrows in dividing cells. Conversely, some intestinal epithelial, spleen, and dorsal root ganglia interphase cells express only CMIIA, organized primarily in networks. Of these, intestinal epithelial cells express both CMIIA and CMIIB when they divide, whereas some dividing cells from both spleen and dorsal root ganglia express only CMIIA and concentrate it in their cleavage furrows. These results suggest that within a given tissue, different cell types express different isoforms of CMII, and that cells expressing either CMIIA or CMIIB alone, or simultaneously, can form a cleavage furrow and divide.

  9. Cytoplasmic tail domain of glycoprotein B is essential for HHV-6 infection.

    Science.gov (United States)

    Mahmoud, Nora F; Jasirwan, Chyntia; Kanemoto, Satoshi; Wakata, Aika; Wang, Bochao; Hata, Yuuki; Nagamata, Satoshi; Kawabata, Akiko; Tang, Huamin; Mori, Yasuko

    2016-03-01

    Human herpesvirus 6 (HHV-6) glycoprotein B (gB) is an abundantly expressed viral glycoprotein required for viral entry and cell fusion, and is highly conserved among herpesviruses. The present study examined the function of HHV-6 gB cytoplasmic tail domain (CTD). A gB CTD deletion mutant was constructed which, in contrast to its revertant, could not be reconstituted. Moreover, deletion of gB cytoplasmic tail impaired the intracellular transport of gB protein to the trans-Golgi network (TGN). Taken together, these results suggest that gB CTD is critical for HHV-6 propagation and important for intracellular transportation. PMID:26802210

  10. Role of "active" potassium transport in the regulation of cytoplasmic pH by nonanimal cells.

    OpenAIRE

    Blatt, M.R.; Slayman, C L

    1987-01-01

    High-affinity potassium uptake in Neurospora occurs by symport with protons [Km (apparent) = 15 microM at pH 5.8], for which a large inward gradient (approximately 400 mV) is generated by the H+-extruding ATPase of the plasma membrane. Operating in parallel, the two transport systems yield a net 1:1 exchange of K+ for cytoplasmic H+. Since this exchange could play a role in cytoplasmic pH (pHi) regulation, the coordinated functioning of the K+-H+ symport and H+ pump has been examined during a...

  11. Perturbation of nucleo-cytoplasmic transport affects size of nucleus and nucleolus in human cells.

    Science.gov (United States)

    Ganguly, Abira; Bhattacharjee, Chumki; Bhave, Madhura; Kailaje, Vaishali; Jain, Bhawik K; Sengupta, Isha; Rangarajan, Annapoorni; Bhattacharyya, Dibyendu

    2016-03-01

    Size regulation of human cell nucleus and nucleolus are poorly understood subjects. 3D reconstruction of live image shows that the karyoplasmic ratio (KR) increases by 30-80% in transformed cell lines compared to their immortalized counterpart. The attenuation of nucleo-cytoplasmic transport causes the KR value to increase by 30-50% in immortalized cell lines. Nucleolus volumes are significantly increased in transformed cell lines and the attenuation of nucleo-cytoplasmic transport causes a significant increase in the nucleolus volume of immortalized cell lines. A cytosol and nuclear fraction swapping experiment emphasizes the potential role of unknown cytosolic factors in nuclear and nucleolar size regulation.

  12. Cytoplasmic proteins of Streptococcus mutans (serotype c) and their interaction with fluoride.

    OpenAIRE

    Yotis, W. W.; Zeb, M; Kubak, B; Wu-Yuan, C; Yotis, E; McNulty, J

    1984-01-01

    The protein profile of the cytoplasmic proteins of Streptococcus mutans GS-5 was determined by two-dimensional gel electrophoresis. Use of this recently developed, high-resolution analytical tool showed in excess of 140 cytoplasmic proteins. The profile consisted of mostly acidic components with pI values between 3.70 and 5.30 and relative molecular weights mainly in the 13,000 to 90,000 range. With sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the proteins were resolved into 40 ...

  13. The early noncoding region of human papillomavirus type 16 is regulated by cytoplasmic polyadenylation factors

    DEFF Research Database (Denmark)

    Glahder, Jacob-Andreas Harald; Kristiansen, Karen; Durand, Marjorie;

    2010-01-01

    All human papillomavirus type 16 (HPV-16) early mRNAs are polyadenylated at the poly(A) signal within the early 3' untranslated region (3'UTR). The 3'end of the early E5 open reading frame and the 3'UTR of HPV-16 is very AU-rich, with five regions similar to cytoplasmic polyadenylation elements...... of the reporter construct containing the HPV-16 early sequences. This repression can be counteracted by a human cytoplasmic poly(A) polymerase, hGLD-2 fused to CPEB1. The hGLD-2/CPEB1 fusion protein facilitates furthermore poly(A) elongation of early HPV transcripts....

  14. Beyond the cytoplasm of Escherichia coli: localizing recombinant proteins where you want them.

    Science.gov (United States)

    Boock, Jason T; Waraho-Zhmayev, Dujduan; Mizrachi, Dario; DeLisa, Matthew P

    2015-01-01

    Recombinant protein expression in Escherichia coli represents a cornerstone of the biotechnology enterprise. While cytoplasmic expression in this host has received the most attention, achieving substantial yields of correctly folded proteins in this compartment can sometimes be met with difficulties. These issues can often be overcome by targeting protein expression to extracytoplasmic compartments (e.g., membrane, periplasm) or to the culture medium. This chapter discusses various strategies for exporting proteins out of the cytoplasm as well as tools for monitoring and optimizing these different export mechanisms. PMID:25447860

  15. Speciation of mercury in salmon egg cell cytoplasm in relation with metallomics research.

    Science.gov (United States)

    Hasegawa, Takuya; Asano, Motoki; Takatani, Kohei; Matsuura, Hirotaka; Umemura, Tomonari; Haraguchi, Hiroki

    2005-12-15

    Speciation of mercury in salmon egg cell cytoplasm was investigated by surfactant-mediated high-performance liquid chromatography/inductively coupled plasma mass spectrometry (HPLC/ICP-MS), where an ODS (octadecylsilica) column coated with a bile acid derivative, CHAPS (3-[(3-cholamidopropyl)-dimethylammonio]-1-propane sulfonate), was used for species separation. Prior to the speciation analysis, total Hg in the cell cytoplasm was determined by ICP-MS at m/z 202 in a flow injection mode. For the precise measurement, salmon egg cell cytoplasm was diluted five-fold with 0.1M Tris (Tris(hydroxymethyl)aminomethane)-HNO(3) buffer solution, and the standard addition method was employed. Thus, the total concentration of Hg in cell cytoplasm was estimated to be 12.4ngg(-1) on the wet weight basis. Next, the cell cytoplasm diluted five-fold with 0.1M Tris-HNO(3) buffer solution was analyzed by surfactant-mediated HPLC with the dual detection system of a UV absorption detector and an ICP-MS instrument. Two peaks corresponding to some proteins and small molecules were mainly observed in those chromatograms. When salmon egg cell cytoplasm was diluted five-fold with 0.01M Tris buffer solution or pure water, some precipitates appeared probably because of precipitation of hydrophobic proteins in cytoplasm. After the precipitates were eliminated with a membrane filter, the filtrate was subjected to the analysis by surfactant-mediated HPLC/UV/ICP-MS. As a result, the peaks for small molecular species of Hg were clearly observed at the retention time near 4.0min (corresponding to low-molecular weight zone) in the chromatograms with UV absorption detection as well as with Hg- and S-specific ICP-MS detections. The small molecule bound with Hg was identified as cysteine through the cysteine-spiked experiment. In addition, the protein fraction on the chromatogram obtained by using the CHAPS-coated ODS column was further analyzed by SEC (size exclusion chromatography). Consequently

  16. Modeling the early steps of cytoplasmic trafficking in viral infection and gene delivery

    OpenAIRE

    Amoruso, Carlo; Lagache, Thibault; Holcman, David

    2011-01-01

    Gene delivery of nucleic acid to the cell nucleus is a fundamental step in gene therapy. In this review of modeling drug and gene delivery, we focus on the particular stage of plasmid DNA or virus cytoplasmic trafficking. A challenging problem is to quantify the success of this limiting stage. We present some models and simulations of plasmid trafficking and of the limiting phase of DNA-polycation escape from an endosome and discuss virus cytoplasmic trafficking. The models can be used to ass...

  17. Paraneoplastic cerebellar degeneration with anti-Yo antibodies - a review.

    Science.gov (United States)

    Venkatraman, Anand; Opal, Puneet

    2016-08-01

    The ataxic syndrome associated with Anti-Yo antibody, or Purkinje cell cytoplasmic antibody type 1 (PCA1), is the most common variant of paraneoplastic cerebellar degeneration (PCD). The typical presentation involves the subacute development of pancerebellar deficits with a clinical plateau within 6 months. The vast majority of cases have been reported in women with pelvic or breast tumors. Magnetic resonance imaging of the brain is often normal in the early stages, with cerebellar atrophy seen later. The underlying mechanism is believed to be an immunological reaction to cerebellar degeneration-related protein 2 (CDR2), a protein usually found in the cerebellum that is ectopically produced by tumor cells. Although both B- and T-cell abnormalities are seen, there is debate about the relative importance of the autoantibodies and cytotoxic T lymphocytes in the neuronal loss. Cerebrospinal fluid abnormalities, primarily elevated protein, lymphocytic pleocytosis, and oligoclonal bands, are common in the early stages. The low prevalence of this condition has not allowed for large-scale randomized controlled trials. Immunotherapies, such as steroids, intravenous immune globulins, and plasma exchange, have been extensively used in managing this condition, with limited success. Although some reports indicate benefit from antitumor therapies like surgery and chemotherapy, this has not been consistently observed. The prognosis for anti-Yo PCD is almost uniformly poor, with most patients left bedridden. Further studies are required to clarify the pathophysiology and provide evidence-based treatment options. PMID:27606347

  18. Paraneoplastic cerebellar degeneration with anti-Yo antibodies - a review.

    Science.gov (United States)

    Venkatraman, Anand; Opal, Puneet

    2016-08-01

    The ataxic syndrome associated with Anti-Yo antibody, or Purkinje cell cytoplasmic antibody type 1 (PCA1), is the most common variant of paraneoplastic cerebellar degeneration (PCD). The typical presentation involves the subacute development of pancerebellar deficits with a clinical plateau within 6 months. The vast majority of cases have been reported in women with pelvic or breast tumors. Magnetic resonance imaging of the brain is often normal in the early stages, with cerebellar atrophy seen later. The underlying mechanism is believed to be an immunological reaction to cerebellar degeneration-related protein 2 (CDR2), a protein usually found in the cerebellum that is ectopically produced by tumor cells. Although both B- and T-cell abnormalities are seen, there is debate about the relative importance of the autoantibodies and cytotoxic T lymphocytes in the neuronal loss. Cerebrospinal fluid abnormalities, primarily elevated protein, lymphocytic pleocytosis, and oligoclonal bands, are common in the early stages. The low prevalence of this condition has not allowed for large-scale randomized controlled trials. Immunotherapies, such as steroids, intravenous immune globulins, and plasma exchange, have been extensively used in managing this condition, with limited success. Although some reports indicate benefit from antitumor therapies like surgery and chemotherapy, this has not been consistently observed. The prognosis for anti-Yo PCD is almost uniformly poor, with most patients left bedridden. Further studies are required to clarify the pathophysiology and provide evidence-based treatment options.

  19. A simple vector system to improve performance and utilisation of recombinant antibodies

    Directory of Open Access Journals (Sweden)

    Vincent Karen J

    2006-12-01

    Full Text Available Abstract Background Isolation of recombinant antibody fragments from antibody libraries is well established using technologies such as phage display. Phage display vectors are ideal for efficient display of antibody fragments on the surface of bacteriophage particles. However, they are often inefficient for expression of soluble antibody fragments, and sub-cloning of selected antibody populations into dedicated soluble antibody fragment expression vectors can enhance expression. Results We have developed a simple vector system for expression, dimerisation and detection of recombinant antibody fragments in the form of single chain Fvs (scFvs. Expression is driven by the T7 RNA polymerase promoter in conjunction with the inducible lysogen strain BL21 (DE3. The system is compatible with a simple auto-induction culture system for scFv production. As an alternative to periplasmic expression, expression directly in the cytoplasm of a mutant strain with a more oxidising cytoplasmic environment (Origami 2™ (DE3 was investigated and found to be inferior to periplasmic expression in BL21 (DE3 cells. The effect on yield and binding activity of fusing scFvs to the N terminus of maltose binding protein (a solubility enhancing partner, bacterial alkaline phosphatase (a naturally dimeric enzymatic reporter molecule, or the addition of a free C-terminal cysteine was determined. Fusion of scFvs to the N-terminus of maltose binding protein increased scFv yield but binding activity of the scFv was compromised. In contrast, fusion to the N-terminus of bacterial alkaline phosphatase led to an improved performance. Alkaline phosphatase provides a convenient tag allowing direct enzymatic detection of scFv fusions within crude extracts without the need for secondary reagents. Alkaline phosphatase also drives dimerisation of the scFv leading to an improvement in performance compared to monovalent constructs. This is illustrated by ELISA, western blot and

  20. Dendritic cell and macrophage staining by monoclonal antibodies in tissue sections and epidermal sheets.

    OpenAIRE

    Flotte, T. J.; Springer, T A; Thorbecke, G. J.

    1983-01-01

    Mouse tissue sections were stained by monoclonal antibodies to macrophage antigens (Mac-1 (M1/70), Mac-2 (M3/38), Mac-3 (M3/84) with the use of immunoperoxidase. Mac-1 was located diffusely in the cytoplasm of round cells in a high percentage of alveolar macrophages, resident peritoneal and bone marrow cells, in splenic red pulp, and in rare perivascular cells in the thymus. Mac-1 was absent in epithelial cells and Langerhans cells. Mac-2 was strongly positive in many dendritic cells in the t...

  1. Regulation of Adherence and Virulence by the Entamoeba histolytica Lectin Cytoplasmic Domain, Which Contains a β2 Integrin Motif

    OpenAIRE

    Vines, Richard R.; Ramakrishnan, Girija; Rogers, Joshua B.; Lockhart, Lauren A.; Mann, Barbara J.; Petri, William A.

    1998-01-01

    Killing of human cells by the parasite Entamoeba histolytica requires adherence via an amebic cell surface lectin. Lectin activity in the parasite is regulated by inside-out signaling. The lectin cytoplasmic domain has sequence identity with a region of the β2 integrin cytoplasmic tail implicated in regulation of integrin-mediated adhesion. Intracellular expression of a fusion protein containing the cytoplasmic domain of the lectin has a dominant negative effect on extracellular lectin-mediat...

  2. Functional effects of anticardiolipin antibodies.

    Science.gov (United States)

    Harris, E N; Pierangeli, S S

    1996-10-01

    The 'lupus anticoagulant' phenomenon is the best documented functional effect of antiphospholipid (aPL) antibodies, occurring either by inhibition of the prothrombinase and/or Factor X activation reactions. Understanding the mechanism by which aPL antibodies inhibit phospholipid dependent coagulation reactions may yield important clues about their 'thrombogenic effects' in vivo. We conducted a series of studies to determine the specificity, diversity, and mechanism by which aPL antibodies inhibit phospholipid dependent reactions. Results showed that purified immunoglobulins with lupus anticoagulant and anti-cardiolipin activities were absorbed by negatively charged phospholipids and both activities were recovered from the phospholipid-antibody precipitate. Purified aPL antibodies inhibited the prothrombinase reaction in a plasma free system in which beta 2-glycoprotein 1 (beta 2-GP1) was absent. Affinity purified aPL antibodies had 25-50 times the inhibitory activity of immunoglobulin preparations. The phospholipid binding proteins, beta 2-GPI and placental anticoagulant protein I (PAP I), independently inhibited the prothrombinase reaction, and when these proteins were combined with aPL, inhibition of the prothrombinase reaction was additive. Antibodies of syphilis had no inhibitory effect, partially accounted for by lack of specificity for phosphotidylserine (PS). Although aPL antibodies inhibited the protein C activation reaction, there was no correlation of these activities with inhibition of the prothrombinase reaction. Together, these results show that aPL exert their effects by interaction with negatively charged phospholipids, in particular phosphotidylserine, but lack of correlation between inhibition of the prothrombinase and protein C activation reactions, suggests that the nature of the coagulation protein is also important. PMID:8902763

  3. Cytological study of radiation induced alterations in cytoplasmic factors controlling male sterility in corn. Progress report, February 28, 1975--December 1, 1975

    International Nuclear Information System (INIS)

    Progress is reported on the following research projects: cytoplasmic constituents of the embryo of various gymnosperms and angiosperms; cytoplasmic male sterility in corn; modification of cytoplasmic sterility factors using gamma radiation, EMS, and ethidium bromide; selection for sterile, blight-resistant corn plants; electron microscopy study of abnormal mitochondria in cytoplasm of corn; cytoplasmic male sterility in Petunia; non-Mendelian variegation in Petunia and Nicotiana; graft transmission of cytoplasmic male sterility; cytoplasmic male sterility in Vicia faba; and studies on Blakeslee's I virus in Datura

  4. Antibodies to watch in 2014.

    Science.gov (United States)

    Reichert, Janice M

    2014-01-01

    Since 2010, mAbs has documented the biopharmaceutical industry's progress in transitioning antibody therapeutics to first Phase 3 clinical studies and regulatory review, and its success at gaining first marketing approvals for antibody-based products. This installment of the "Antibodies to watch" series outlines events anticipated to occur between December 2013 and the end of 2014, including first regulatory actions on marketing applications for vedolizumab, siltuximab, and ramucirumab, as well as the Fc fusion proteins Factor IX-Fc and Factor VIII-Fc; and the submission of first marketing applications for up to five therapeutics (secukinumab, ch14.18, onartuzumab, necitumumab, gevokizumab). Antibody therapeutics in Phase 3 studies are described, with an emphasis on those with study completion dates in 2014, including antibodies targeting interleukin-17a or the interleukin-17a receptor (secukinumab, ixekizumab, brodalumab), proprotein convertase subtilisin/kexin type 9 (alirocumab, evolocumab, bococizumab), and programmed death 1 receptor (lambrolizumab, nivolumab). Five antibodies with US Food and Drug Administration's Breakthrough Therapy designation (obinutuzumab, ofatumumab, lambrolizumab, bimagrumab, daratumumab) are also discussed. PMID:24284914

  5. Radiolabeled monoclonal antibodies: a review

    International Nuclear Information System (INIS)

    Since the description by Kohler and Milstein 1975 of their technique for producing monoclonal antibodies of predefined specificity, it has become a mainstay in most laboratories that utilize immunochemical techniques to study problems in basic, applied or clinical research. Paradoxically, the very success of monoclonal antibodies has generated a literature which is now so vast and scattered that it has become difficult to obtain a perspective. This brief review represents the distillation of many publications relating to the production and use of monoclonaal antibodies as radiopharmaceuticals. Significant advances were made possible in the last few years by combined developments in the fields of tumor-associated antigens and of monoclonal antibodies. In fact monoclonal antibodies against some well defined tumor-associated antigens, has led to significantly greater practical possibilities for producing highly specific radiolabeled antibodies as radiopharmaceuticals for diagnosis and therapy of human tumors. One of the main requirements of this methodology is the availability of stable radiopharmaceutical reagents which after labeling in vivo injection retain the capacity of specific interaction with the defined antigen and their molecular integrity. Since injection into human is the objetive of this kind of study all the specifications of radiopharmaceutical have to be fulfilled e.g. sterility, apirogenicity and absence of toxicity. (author)

  6. Radioimmunoguided surgery using monoclonal antibody

    International Nuclear Information System (INIS)

    The potential proficiency of radioimmunoguided surgery in the intraoperative detection of tumors was assessed using labeled monoclonal antibody B72.3 in 66 patients with tissue-proved tumor. Monoclonal antibody B72.3 was injected 5 to 42 days preoperatively, and the hand-held gamma-detecting probe was used intraoperatively to detect the presence of tumor. Intraoperative probe counts of less than 20 every 2 seconds, or tumor-to-adjacent normal tissue ratios less than 2:1 were considered negative (system failure). Positive probe counts were detected in 5 of 6 patients with primary colon cancer (83 percent), in 31 of 39 patients with recurrent colon cancer (79 percent), in 4 of 5 patients with gastric cancer (80 percent), in 3 of 8 patients with breast cancer (37.5 percent), and in 4 of 8 patients with ovarian cancer (50 percent) undergoing second-look procedures. Additional patients in each group were scored as borderline positive. Overall, radioimmunoguided surgery using B72.3 identified tumors in 47 patients (71.2 percent), bordered on positive in 6 patients (9.1 percent), and failed to identify tumor in 13 patients (19.7 percent). Improved selection of patients for antigen-positive tumors, the use of higher affinity second-generation antibodies, alternate routes of antibody administration, alternate radionuclides, and more sophisticatedly bioengineered antibodies and antibody combinations should all lead to improvements in radioimmunoguided surgery

  7. Tabhu: tools for antibody humanization.

    KAUST Repository

    Olimpieri, Pier Paolo

    2014-10-09

    SUMMARY: Antibodies are rapidly becoming essential tools in the clinical practice, given their ability to recognize their cognate antigens with high specificity and affinity, and a high yield at reasonable costs in model animals. Unfortunately, when administered to human patients, xenogeneic antibodies can elicit unwanted and dangerous immunogenic responses. Antibody humanization methods are designed to produce molecules with a better safety profile still maintaining their ability to bind the antigen. This can be accomplished by grafting the non-human regions determining the antigen specificity into a suitable human template. Unfortunately, this procedure may results in a partial or complete loss of affinity of the grafted molecule that can be restored by back-mutating some of the residues of human origin to the corresponding murine ones. This trial-and-error procedure is hard and involves expensive and time-consuming experiments. Here we present tools for antibody humanization (Tabhu) a web server for antibody humanization. Tabhu includes tools for human template selection, grafting, back-mutation evaluation, antibody modelling and structural analysis, helping the user in all the critical steps of the humanization experiment protocol. AVAILABILITY: http://www.biocomputing.it/tabhu CONTACT: anna.tramontano@uniroma1.it, pierpaolo.olimpieri@uniroma1.it SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

  8. Avian Diagnostic and Therapeutic Antibodies

    Energy Technology Data Exchange (ETDEWEB)

    Bradley, David Sherman [UND SMHS

    2012-12-31

    A number of infectious agents have the potential of causing significant clinical symptomology and even death, but dispite this, the number of incidence remain below the level that supports producing a vaccine. Therapeutic antibodies provide a viable treatment option for many of these diseases. We proposed that antibodies derived from West Nile Virus (WNV) immunized geese would be able to treat WNV infection in mammals and potential humans. We demonstrated that WNV specific goose antibodies are indeed successful in treating WNV infection both prophylactically and therapeutically in a golden hamster model. We demonstrated that the goose derived antibodies are non-reactogenic, i.e. do not cause an inflammatory response with multiple exposures in mammals. We also developed both a specific pathogen free facility to house the geese during the antibody production phase and a patent-pending purification process to purify the antibodies to greater than 99% purity. Therefore, the success of these study will allow a cost effective rapidly producible therapeutic toward clinical testing with the necessary infrastructure and processes developed and in place.

  9. Production and characterization of peptide antibodies

    DEFF Research Database (Denmark)

    Trier, Nicole Hartwig; Hansen, Paul Robert; Houen, Gunnar

    2012-01-01

    Proteins are effective immunogens for generation of antibodies. However, occasionally the native protein is known but not available for antibody production. In such cases synthetic peptides derived from the native protein are good alternatives for antibody production. These peptide antibodies are...

  10. Primary structure and functional scFv antibody expression of an antibody against the human protooncogen c-myc.

    Science.gov (United States)

    Fuchs, P; Breitling, F; Little, M; Dübel, S

    1997-06-01

    The immunoglobulin heavy- and light-chain variable region (Vh and Vl) genes were isolated from Myc1-9E10 hybridoma cells, which secreted monoclonal antibody against human oncogen c-myc. The expression vector pOPE52-c-myc was constructed for the recombinant production in E. coli. A 30 kDa single chain fragment (scFv) expression product was found in the periplasmic space by SDS-PAGE and immunoblotting. A significant fraction was processed correctly as demonstrated with an antiserum recognizing the processed aminoterminus only. The specific binding of the scFv fragment to the peptide epitope of the maternal monoclonal antibody was demonstrated and the primary sequence of the variable regions was determined. Sequence comparison with previously published partial Vh and Vl sequences from this hybridoma cell line revealed a genetic heterogeneity for the light chain variable region. The potential use of this scFv as a new tool for detection and purification of tagged proteins, for adding costimulatory signals to the surface of cancer cells as well as for analyzing c-myc function in the living cell by cytoplasmic expression is discussed.

  11. CONTINUOUS MEASUREMENT OF THE CYTOPLASMIC PH IN LACTOCOCCUS-LACTIS WITH A FLUORESCENT PH INDICATOR

    NARCIS (Netherlands)

    MOLENAAR, D; ABEE, T; KONINGS, WN

    1991-01-01

    The cytoplasmic pH of Lactococcus lactis was studied with the fluorescent pH indicator 2',7'-bis-(2-carboxyethyl)-5 (and-6)-carboxyfluorescein (BCECF). A novel method was applied for loading bacterial cells with BCECF, which consists of briefly treating a dense cell suspension with acid in the prese

  12. Ubiquitin-dependent distribution of the transcriptional coactivator p300 in cytoplasmic inclusion bodies.

    Science.gov (United States)

    Chen, Jihong; Halappanavar, Sabina; Th' ng, John P H; Li, Qiao

    2007-01-01

    The protein level of transcriptional coactivator p300, an essential nuclear protein, is critical to a broad array of cellular activities including embryonic development, cell differentiation and proliferation. We have previously established that histone deacetylase inhibitor such as valproic acid induces p300 degradation through the 26S proteasome pathway. Here, we report the roles of cellular trafficking and spatial redistribution in valproic acid-induced p300 turnover. Our study demonstrates that p300 is redistributed to the cytoplasm prior to valproic acid-induced turnover. Inhibition of proteasome-dependent protein degradation, does not prevent nucleo-cytoplasmic shuttling of p300, rather sequesters the cytoplasmic p300 to a distinct perinuclear region. In addition, the formation of p300 aggregates in the perinuclear region depends on functional microtubule networks and correlates with p300 ubiquitination. Our work establishes, for the first time, that p300 is also a substrate of the cytoplasmic ubiquitin-proteasome system and provides insight on how cellular trafficking and spatial redistribution regulate the availability and activity of transcriptional coactivator p300.

  13. Localization and topogenesis studies of cytoplasmic and vacuolar homologs of the Galanthus nivalis agglutinin.

    Science.gov (United States)

    Fouquaert, Elke; Hanton, Sally L; Brandizzi, Federica; Peumans, Willy J; Van Damme, Els J M

    2007-07-01

    The Galanthus nivalis agglutinin (GNA) is synthesized as a preproprotein. To corroborate the role of the different targeting peptides in the topogenesis of GNA and related proteins, different constructs were made whereby both the complete original GNA gene and different truncated sequences were coupled to the enhanced green fluorescent protein (EGFP). In addition, a GNA ortholog from rice that lacks the signal peptide and C-terminal propeptide sequence was fused to EGFP. These fusion constructs were expressed in tobacco BY-2 cells and their localization analyzed by confocal fluorescence microscopy. We observed that the processed preproprotein of GNA was directed towards the vacuolar compartment, whereas both the truncated forms of GNA corresponding to the mature lectin polypeptide and the rice ortholog of GNA were located in the nucleus and the cytoplasm. It can be concluded, therefore, that removal of the C-terminal propeptide and the signal peptide is sufficient to change the subcellular targeting of a normally vacuolar protein to the nuclear/cytoplasmic compartment of the BY-2 cells. These findings support the proposed hypothesis that cytoplasmic/nuclear GNA-like proteins and their vacuolar homologs are evolutionarily related and that the classical GNA-related lectins might have evolved from cytoplasmic orthologs through an evolutionary event involving the insertion of a signal peptide and a C-terminal propeptide.

  14. Probing cytoplasmic organization and the actin cytoskeleton of plant cells with optical tweezers

    NARCIS (Netherlands)

    Ketelaar, T.; Honing, van der H.S.; Emons, A.M.C.

    2010-01-01

    In interphase plant cells, the actin cytoskeleton is essential for intracellular transport and organization. To fully understand how the actin cytoskeleton functions as the structural basis for cytoplasmic organization, both molecular and physical aspects of the actin organization have to be conside

  15. Water- and Air-Distributed Conidia Differ in Sterol Content and Cytoplasmic Microviscosity▿

    OpenAIRE

    van, Leeuwen, M.; Doorn, van, Erik A.; Golovina, E. A.; Stark, J.; Dijksterhuis, J.

    2009-01-01

    Airborne and waterborne fungal spores were compared with respect to cytoplasmic viscosity and the presence of ergosterol. These parameters differed markedly between the two spore types and correlated with spore survival. This suggests that the mode of spore dispersal has a bearing on cellular composition, which is relevant for the eradication of industrially relevant fungal propagules.

  16. Water- and air-distributed conidia differ in sterol content and cytoplasmic microviscosity

    NARCIS (Netherlands)

    Leeuwen, van M.R.; Doorn, van T.M.; Golovina, E.A.; Stark, J.; Dijksterhuis, J.

    2010-01-01

    Airborne and waterborne fungal spores were compared with respect to cytoplasmic viscosity and the presence of ergosterol. These parameters differed markedly between the two spore types and correlated with spore survival. This suggests that the mode of spore dispersal has a bearing on cellular compos

  17. The nuclear protein Sam68 is recruited to the cytoplasmic stress granules during enterovirus 71 infection.

    Science.gov (United States)

    Zhang, Hua; Chen, Ning; Li, Pengfei; Pan, Ziye; Ding, Yun; Zou, Dehua; Li, Liyang; Xiao, Lijie; Shen, Binglei; Liu, Shuxia; Cao, Hongwei; Cui, Yudong

    2016-07-01

    Our previous study found that the nuclear protein, 68-kDa Src-associated in mitosis protein (Sam68), is translocated to the cytoplasm and forms punctate pattern during enterovirus 71 (EV71) infection [Virus Research, 180 (2014), 1-11]. However, the exact function of this punctate pattern in cytoplasm during EV71 infection remains unknown. In this study, we firstly have examined this punctate pattern of Sam68 re-localization in the cytoplasm, and observed the obvious recruitments of Sam68 to the EV71-induced stress granules (SGs). Sam68, belongs to the KH domain family of RNA binding proteins (RBPs), was then confirmed that its KH domain was essential for this recruitment. Nevertheless, Knockdown of Sam68 expression using ShRNA had no effects on SGs assembly, indicating that Sam68 is not a constitutive component of the SGs during EV71 infection. Lastly, we investigated the importance of microtubulin transport to SGs aggregation, and revealed that microtubule depolymerization inhibited SGs formation, suggesting that EV71-induced SGs move throughout the cytoplasm in a microtubule-dependent manner. Taken together, these results illuminated that EV71 infections can induce SGs formation, and Sam68, as a SGs component, migrates alone with SGs dependent on intact microtubule upon the viral infections. These findings may provide novel underlying mechanism for delineating the role of SGs during EV71 infection. PMID:27057671

  18. Guanidino groups greatly enhance the action of antimicrobial peptidomimetics against bacterial cytoplasmic membranes

    DEFF Research Database (Denmark)

    Andreev, Konstantin; Bianchi, Christopher; Laursen, Jonas Striegler;

    2014-01-01

    monolayers composed of 1,2-dipalmitoyl-sn-glycero-3-phosphatidylglycerol (DPPG) were used to model cytoplasmic membranes of both Gram-positive and Gram-negative bacteria, while lipopolysaccharide Kdo2-lipid A monolayers were mimicking the outer membrane of Gram-negative species. We report the results...

  19. Cytoplasmic transfer of oligomycin resistance during protoplast fusion of Saccharomycopsis lipolytica.

    OpenAIRE

    Matsuoka, M; Uchida, K.(Physikalisches Institut, University of Bonn, Bonn, Germany); Aiba, S

    1982-01-01

    Mitotic segregation of oligomycin resistance and oligomycin sensitivity was observed among the prototrophic progeny of protoplast fusion between drug-resistant and drug-sensitive complementary auxotrophs of Saccharomycopsis lipolytica. The transfer of oligomycin resistance by protoplast fusion without karyogamy suggests a cytoplasmic inheritance of this drug resistance determinant.

  20. Mitochondrial DNA variation within P-type cytoplasmic male sterility of Plantago lanceolata L

    NARCIS (Netherlands)

    Groenendijk, C.F.M.; Sandbrink, J.M.; Van Brederode, J.; Van Damme, J.M.M.

    1997-01-01

    MtDNA restriction fragment polymorphisms were found between cytoplasmic male-sterility types P and R of Plantago lanceolata with the homologous probe pPl311 and maize mtDNA fragments derived from the regions of atp1, cox1 and cox2. No mtDNA differences were observed between male-sterile and restored

  1. Cytoplasmic male sterility in Petunia hybrida. A structural and histochemical analysis.

    NARCIS (Netherlands)

    Bino, R.J.

    1986-01-01

    This thesis presents an analysis of the structural and histochemical aspects of cytoplasmic male sterility (cms) in Petuniahybrida . In petunia and in other crops, cms is the most commonly used tool for hybrid seed production. Application of the trait makes hybrid seed production possi

  2. Establishment and Identification of Cytoplasmic Male Sterility in Brassica napus by Intergeneric Somatic Hybridization

    Institute of Scientific and Technical Information of China (English)

    HU Qiong; LI Yun-chang; MEI De-sheng; FANG Xiao-ping; Lise N Hansen; Sven B Andersen

    2003-01-01

    Exploitation of novel cytoplasmic male sterility(CMS)is a main approach for widening the cytoplasmic genetic background of hybrid oilseed rape and avoiding epidemic risk in oilseed rape production.In this study,symmetric somatic hybrids between Brassicanapus var.Zhongshuang4 and Sinapis arvensis(Yeyou18)were produced by protoplast fusion.Two of the six established hybrids were male sterile showing trace or no pollen release upon flowering with non-or slightly extended stamens.Using Zhongshuang4 as a recurrent parent to pollinate the male sterile plants,the ratio of male sterile plants increased with the number of backcrosses.As early as in BC3 generation,most of the sterile families had nearly 100%sterile plants.Up to BC4 generation,the male sterility became stable and no fertility segregation was observed.All F1 progenies from tested crosses using restorer and maintainer lines of Polima CMS were 100%sterile,indicating that the established CMS by somatic hybridization is different from Polima CMS.The origin of the cytoplasm and potential use of this hovel CMS in oilseed rape breeding were discussed.Key wotds:Oilseed rape,Protoplast fusion,Cytoplasmic male sterility,Sinapis arvensis

  3. Cytoplasmic male sterility and inter and intra subgenomic heterosis studies in Brassica species: A review

    Directory of Open Access Journals (Sweden)

    Rameeh Valiollah

    2014-01-01

    Full Text Available Plants of the genus Brassica comprise a remarkably diverse group of crops and encompass varieties that are grown as oilseeds, vegetables, condiment mustards and forages. One of the basic requirements for developing hybrid varieties in oilseed Brassica is the availability of proven heterosis. The development of hybrid cultivars has been successful in many Brassica spp. Midparent heterosis and high-parent heterosis (heterobeltiosis have extensively been explored and utilized for boosting various quantity and quality traits in rapeseed. Heterosis is commercially exploited in rapeseed and its potential use has been demonstrated in turnip rape (B. rapa L. and Indian mustard (B. juncea L. for seed yield and most of the agronomic traits. The oilseed rape plant, B. napus, possesses two endogenous male sterile cytoplasms, nap and pol. Ogura type of cytoplasmic male sterility was first discovered in Japanese wild radish and other male-sterile Brassicas (Ogura bearing cytoplasm derived from interspecific crosses. Information concerning the allelic frequencies of restorers can be useful in trying to understand their evolutionary origins. The ogu, pol and nap cytoplasms of B. napus induce sterility in all, some, and only a few cultivars, respectively. In this study, different kinds of male sterility, combining ability and heterosis of qualitative and quantitative traits in different Brassica species will be reviеwed.

  4. Molecular characterization of a new type of cytoplasmic male sterile sugar beet

    Institute of Scientific and Technical Information of China (English)

    谢纬武; 康传红; 王继志; 王斌; 郭德栋

    1996-01-01

    A line (named Cl) of cytoplasmic sterility of sugar beet whose cytoplasm derived from Betacicla Turkey was obtained by interspecific hybrid. Its cytoplasm and a spontaneous male sterile cytoplasm from wild beet Beta maritima (named M) were compared with that of Owen’s sterile line (S-cms) and a common maintainer of them named N was used as control. RFLP and RAPD methods were mainly used in our experiments. The restriction fragment patterns of mtDNAs were found to be likely but for a few of specific low-lighted electrophoresis bands in Cl. The results of Southern hybridization of six heterogeneous mitochondrial genes as probes to digests of mtDNAs by six restriction enzymes showed to be analogous between S and M lines. But the Cl mtDNA was sorted out by hybridization of atpA probe. Difference of low-molecular-weight mitochondrial DNAs was found among the three sterile lines. Three RNA molecules weighing about 4.2kb stably existed in Cl mitochondria. Our results of RAPD also supported that the Cl cytoplas

  5. Transport of the outer dynein arm complex to cilia requires a cytoplasmic protein Lrrc6.

    Science.gov (United States)

    Inaba, Yasuko; Shinohara, Kyosuke; Botilde, Yanick; Nabeshima, Ryo; Takaoka, Katsuyoshi; Ajima, Rieko; Lamri, Lynda; Takeda, Hiroyuki; Saga, Yumiko; Nakamura, Tetsuya; Hamada, Hiroshi

    2016-07-01

    Lrrc6 encodes a cytoplasmic protein that is expressed specifically in cells with motile cilia including the node, trachea and testes of the mice. A mutation of Lrrc6 has been identified in human patients with primary ciliary dyskinesia (PCD). Mutant mice lacking Lrrc6 show typical PCD defects such as hydrocephalus and laterality defects. We found that in the absence of Lrrc6, the morphology of motile cilia remained normal, but their motility was completely lost. The 9 + 2 arrangement of microtubules remained normal in Lrrc6(-/-) mice, but the outer dynein arms (ODAs), the structures essential for the ciliary beating, were absent from the cilia. In the absence of Lrrc6, ODA proteins such as DNAH5, DNAH9 and IC2, which are assembled in the cytoplasm and transported to the ciliary axoneme, remained in the cytoplasm and were not transported to the ciliary axoneme. The IC2-IC1 interaction, which is the first step of ODA assembly, was normal in Lrrc6(-/-) mice testes. Our results suggest that ODA proteins may be transported from the cytoplasm to the cilia by an Lrrc6-dependent mechanism.

  6. Cytoplasmic membrane changes during adaptation of the fresh water cyanobacterium Synechococcus 6311 to salinity

    Science.gov (United States)

    Lefort-Tran, M.; Pouphile, M.; Spath, S.; Packer, L.

    1988-01-01

    In this investigation, changes were characterized in cell structure and cytoplasmic membrane organization that occur when the freshwater cyanobacterium Synechococcus 6311 is transferred from 'low salt' (0.03 molar NaCl) to 'high salt' (0.5 molar NaCl) media (i.e. sea water concentration). Cells were examined at several time points after the imposition of the salt stress and compared to control cells, in thin sections and freeze fracture electron microscopy, and by flow cytometry. One minute after exposure to high salt, i.e. 'salt shock', virtually all intracellular granules disappeared, the density of the cytoplasm decreased, and the appearance of DNA material was changed. Glycogen and other granules, however, reappeared by 4 hours after salt exposure. The organization of the cytoplasmic membrane undergoes major reorganization following salt shock. Freeze-fracture electron microscopy showed that small intramembrane particles (diameter 7.5 and 8.5 nanometers) are reduced in number by two- to fivefold, whereas large particles, (diameters 14.5 and 17.5 nanometers) increase two- to fourfold in frequency, compared to control cells grown in low salt medium. The changes in particle size distribution suggest synthesis of new membrane proteins, in agreement with the known increases in respiration, cytochrome oxidase, and sodium proton exchange activity of the cytoplasmic membrane.

  7. Cytoplasmic dynein and its regulatory proteins in Golgi pathology in nervous system disorders

    NARCIS (Netherlands)

    Jaarsma, Dick; Hoogenraad, Casper C

    2015-01-01

    The Golgi apparatus is a dynamic organelle involved in processing and sorting of lipids and proteins. In neurons, the Golgi apparatus is important for the development of axons and dendrites and maintenance of their highly complex polarized morphology. The motor protein complex cytoplasmic dynein has

  8. The 75-kilodalton cytoplasmic Chlamydia trachomatis L2 polypeptide is a DnaK-like protein

    DEFF Research Database (Denmark)

    Birkelund, Svend; Lundemose, AG; Christiansen, Gunna

    1990-01-01

    The gene coding for the 75-kilodalton cytoplasmic Chlamydia trachomatis L2 polypeptide has been cloned in Escherichia coli, and the nucleotide sequence has been determined. The cloned DNA fragment contained the coding region as well as the putative promoter. The deduced amino acid sequence of the 1...

  9. A Putative NES Mediates Cytoplasmic Localization of Apoptin in Normal Cells

    Institute of Scientific and Technical Information of China (English)

    Qing-Ming WANG; Guo-Cai FAN; Ji-Zhong CHEN; Hui-Peng CHEN; Fu-Chu HE

    2004-01-01

    Apoptin, a protein expressed by chicken anemia virus, is found predominantly in the cytoplasm in normal cells, whereas it localizes in the nucleus in transformed and malignant cells. However, the mechanisms that regulate the different subcellular localization of Apoptin in normal and tumor cells have not been fully clarified. In this work, a putative nuclear export signal (NES) in Apoptin was predicted. It was testified that the putative NES (pNES) of Apoptin was not a functional NES, but actually acted as a cytoplasmic retention signal. Deletion of the pNES led to the nuclear accumulation of Apoptin in normal cells. In addition,when a strong nuclear localization signal was introduced into Apoptin, it exclusively translocated to the nucleus in normal cells. These observations indicated that the cytoplasmic localization of Apoptin in normal cells results from the balance between cytoplasmic retention and nuclear import. On the other hand, the pNES was also proved to be necessary for Apoptin multimerization. Mutants lacking the pNES did not form obviously visible globular aggregates in normal or tumor cells.

  10. Effects of ozone on apoplast/cytoplasm partitioning of ascorbic acid in snap bean

    Energy Technology Data Exchange (ETDEWEB)

    Burkey, K.O. [North Carolina State Univ., Dept. of Crop Science and Botany, Raleigh, NC (United States)

    1999-07-01

    Apoplast/cytoplasm partitioning of ascorbic acid (AA) was examined in four genotypes of snap bean (Phaseolus vulgaris L.) known to differ in ozone sensitivity. Plants were grown in pots under field conditions using open-top chambers to establish charcoal-filtered (CF) air (36 nmol mol{sup -1} ozone) or elevated ozone (77 nmol mol{sup -1} ozone) treatments, AA in fully expanded leaves of 36-day-old plants was separated into apoplast and cytoplasm fractions by vacuum infiltration methods using glucose 6-phosphate as a marker for cytoplasm contamination, Apoplast ascorbate levels ranged from 30 to 150 nmol g{sup -1} fresh weight. Ozone-sensitive genotypes partitioned 1-2% of total AA into the apoplast under CF conditions and up to 7% following a 7-day ozone exposure. In contrast, an ozone-tolerant genotype partitioned 3-4% of total leaf AA into the leaf apoplast in both CF and ozone-treated plants. The results suggest that genetic background and ozone stress are factors that affect AA levels in the extracellular space. For all genotypes, the fraction of AA in the oxidized form was higher in the apoplast compared to the cytoplasm, indicative of a more oxidizing environment within the cell wall. (au)

  11. Enhanced electroporation in plant tissues via low frequency pulsed electric fields: influence of cytoplasmic streaming.

    Science.gov (United States)

    Asavasanti, Suvaluk; Stroeve, Pieter; Barrett, Diane M; Jernstedt, Judith A; Ristenpart, William D

    2012-01-01

    Pulsed electric fields (PEF) are known to be effective at permeabilizing plant tissues. Prior research has demonstrated that lower pulse frequencies induce higher rates of permeabilization, but the underlying reason for this response is unclear. Intriguingly, recent microscopic observations with onion tissues have also revealed a correlation between PEF frequency and the subsequent speed of intracellular convective motion, i.e., cytoplasmic streaming. In this paper, we investigate the effect of cytoplasmic streaming on the efficacy of plant tissue permeabilization via PEF. Onion tissue samples were treated with Cytochalasin B, a known inhibitor of cytoplasmic streaming, and changes in cellular integrity and viability were measured over a wide range of frequencies and field strengths. We find that at low frequencies (f streaming results in a 19% decrease in the conductivity disintegration index compared with control samples. Qualitatively, similar results were observed using a microscopic cell viability assay. The results suggest that at low frequencies convection plays a statistically significant role in distributing more conductive fluid throughout the tissue, making subsequent pulses more efficacious. The key practical implication is that PEF pretreatment at low frequency can increase the rate of tissue permeabilization in dehydration or extraction processes, and that the treatment will be most effective when cytoplasmic streaming is most active, i.e., with freshly prepared plant tissues. PMID:22246974

  12. Production of recombinant antibodies using bacteriophages

    OpenAIRE

    Shukra, A. M.; Sridevi, N. V.; Dev Chandran,; Kapil Maithal,

    2014-01-01

    Recombinant antibody fragments such as Fab, scFv, diabodies, triabodies, single domain antibodies and minibodies have recently emerged as potential alternatives to monoclonal antibodies, which can be engineered using phage display technology. These antibodies match the strengths of conventionally produced monoclonal antibodies and offer advantages for the development of immunodiagnostic kits and assays. These fragments not only retain the specificity of the whole monoclonal ...

  13. Cytoplasmic CUG RNA foci are insufficient to elicit key DM1 features.

    Directory of Open Access Journals (Sweden)

    Warunee Dansithong

    Full Text Available The genetic basis of myotonic dystrophy type I (DM1 is the expansion of a CTG tract located in the 3' untranslated region of DMPK. Expression of mutant RNAs encoding expanded CUG repeats plays a central role in the development of cardiac disease in DM1. Expanded CUG tracts form both nuclear and cytoplasmic aggregates, yet the relative significance of such aggregates in eliciting DM1 pathology is unclear. To test the pathophysiology of CUG repeat encoding RNAs, we developed and analyzed mice with cardiac-specific expression of a beta-galactosidase cassette in which a (CTG(400 repeat tract was positioned 3' of the termination codon and 5' of the bovine growth hormone polyadenylation signal. In these animals CUG aggregates form exclusively in the cytoplasm of cardiac cells. A key pathological consequence of expanded CUG repeat RNA expression in DM1 is aberrant RNA splicing. Abnormal splicing results from the functional inactivation of MBNL1, which is hypothesized to occur due to MBNL1 sequestration in CUG foci or from elevated levels of CUG-BP1. We therefore tested the ability of cytoplasmic CUG foci to elicit these changes. Aggregation of CUG RNAs within the cytoplasm results both in Mbnl1 sequestration and in approximately a two fold increase in both nuclear and cytoplasmic Cug-bp1 levels. Significantly, despite these changes RNA splice defects were not observed and functional analysis revealed only subtle cardiac dysfunction, characterized by conduction defects that primarily manifest under anesthesia. Using a human myoblast culture system we show that this transgene, when expressed at similar levels to a second transgene, which encodes expanded CTG tracts and facilitates both nuclear focus formation and aberrant splicing, does not elicit aberrant splicing. Thus the lack of toxicity of cytoplasmic CUG foci does not appear to be a consequence of low expression levels. Our results therefore demonstrate that the cellular location of CUG RNA

  14. Antibody-Directed Phototherapy (ADP

    Directory of Open Access Journals (Sweden)

    M. Adil Butt

    2013-04-01

    Full Text Available Photodynamic therapy (PDT is a clinically-approved but rather under-exploited treatment modality for cancer and pre-cancerous superficial lesions. It utilises a cold laser or LED to activate a photochemical reaction between a light activated drug (photosensitiser-drug and oxygen to generate cytotoxic oxygen species. These free radical species damage cellular components leading to cell death. Despite its benefits, the complexity, limited potency and side effects of PDT have led to poor general usage. However, the research area is very active with an increasing understanding of PDT-related cell biology, photophysics and significant progress in molecular targeting of disease. Monoclonal antibody therapy is maturing and the next wave of antibody therapies includes antibody-drug conjugates (ADCs, which promise to be more potent and curable. These developments could lift antibody-directed phototherapy (ADP to success. ADP promises to increase specificity and potency and improve drug pharmacokinetics, thus delivering better PDT drugs whilst retaining its other benefits. Whole antibody conjugates with first generation ADP-drugs displayed problems with aggregation, poor pharmacokinetics and loss of immuno-reactivity. However, these early ADP-drugs still showed improved selectivity and potency. Improved PS-drug chemistry and a variety of conjugation strategies have led to improved ADP-drugs with retained antibody and PS-drug function. More recently, recombinant antibody fragments have been used to deliver ADP-drugs with superior drug loading, more favourable pharmacokinetics, enhanced potency and target cell selectivity. These improvements offer a promise of better quality PDT drugs.

  15. Myosin-Powered Membrane Compartment Drives Cytoplasmic Streaming, Cell Expansion and Plant Development.

    Directory of Open Access Journals (Sweden)

    Valera V Peremyslov

    Full Text Available Using genetic approaches, particle image velocimetry and an inert tracer of cytoplasmic streaming, we have made a mechanistic connection between the motor proteins (myosins XI, cargo transported by these motors (distinct endomembrane compartment defined by membrane-anchored MyoB receptors and the process of cytoplasmic streaming in plant cells. It is shown that the MyoB compartment in Nicotiana benthamiana is highly dynamic moving with the mean velocity of ~3 μm/sec. In contrast, Golgi, mitochondria, peroxisomes, carrier vesicles and a cytosol flow tracer share distinct velocity profile with mean velocities of 0.6-1.5 μm/sec. Dominant negative inhibition of the myosins XI or MyoB receptors using overexpression of the N. benthamiana myosin cargo-binding domain or MyoB myosin-binding domain, respectively, resulted in velocity reduction for not only the MyoB compartment, but also each of the tested organelles, vesicles and cytoplasmic streaming. Furthermore, the extents of this reduction were similar for each of these compartments suggesting that MyoB compartment plays primary role in cytosol dynamics. Using gene knockout analysis in Arabidopsis thaliana, it is demonstrated that inactivation of MyoB1-4 results in reduced velocity of mitochondria implying slower cytoplasmic streaming. It is also shown that myosins XI and MyoB receptors genetically interact to contribute to cell expansion, plant growth, morphogenesis and proper onset of flowering. These results support a model according to which myosin-dependent, MyoB receptor-mediated transport of a specialized membrane compartment that is conserved in all land plants drives cytoplasmic streaming that carries organelles and vesicles and facilitates cell growth and plant development.

  16. Myosin-Powered Membrane Compartment Drives Cytoplasmic Streaming, Cell Expansion and Plant Development

    Science.gov (United States)

    Peremyslov, Valera V.; Cole, Rex A.; Fowler, John E.; Dolja, Valerian V.

    2015-01-01

    Using genetic approaches, particle image velocimetry and an inert tracer of cytoplasmic streaming, we have made a mechanistic connection between the motor proteins (myosins XI), cargo transported by these motors (distinct endomembrane compartment defined by membrane-anchored MyoB receptors) and the process of cytoplasmic streaming in plant cells. It is shown that the MyoB compartment in Nicotiana benthamiana is highly dynamic moving with the mean velocity of ~3 μm/sec. In contrast, Golgi, mitochondria, peroxisomes, carrier vesicles and a cytosol flow tracer share distinct velocity profile with mean velocities of 0.6–1.5 μm/sec. Dominant negative inhibition of the myosins XI or MyoB receptors using overexpression of the N. benthamiana myosin cargo-binding domain or MyoB myosin-binding domain, respectively, resulted in velocity reduction for not only the MyoB compartment, but also each of the tested organelles, vesicles and cytoplasmic streaming. Furthermore, the extents of this reduction were similar for each of these compartments suggesting that MyoB compartment plays primary role in cytosol dynamics. Using gene knockout analysis in Arabidopsis thaliana, it is demonstrated that inactivation of MyoB1-4 results in reduced velocity of mitochondria implying slower cytoplasmic streaming. It is also shown that myosins XI and MyoB receptors genetically interact to contribute to cell expansion, plant growth, morphogenesis and proper onset of flowering. These results support a model according to which myosin-dependent, MyoB receptor-mediated transport of a specialized membrane compartment that is conserved in all land plants drives cytoplasmic streaming that carries organelles and vesicles and facilitates cell growth and plant development. PMID:26426395

  17. Cocaine-associated retiform purpura: a C5b-9-mediated microangiopathy syndrome associated with enhanced apoptosis and high levels of intercellular adhesion molecule-1 expression.

    Science.gov (United States)

    Magro, Cynthia M; Wang, Xuan

    2013-10-01

    Cocaine-associated retiform purpura is a recently described entity characterized by striking hemorrhagic necrosis involving areas of skin associated with administration of cocaine. Levamisole, an adulterant in cocaine, has been suggested as the main culprit pathogenetically. Four cases of cocaine-associated retiform purpura were encountered in the dermatopathology practice of C. M. Magro. The light microscopic findings were correlated with immunohistochemical and immunofluorescence studies. All 4 cases showed a very striking thrombotic diathesis associated with intravascular macrophage accumulation. Necrotizing vasculitis was noted in 1 case. Striking intercellular adhesion molecule-1 (ICAM-1)/CD54 expression in vessel wall along with endothelial expression of caspase 3 and extensive vascular C5b-9 deposition was observed in all biopsies examined. Cocaine-induced retiform purpura is a C5b-9-mediated microvascular injury associated with enhanced apoptosis and prominent vascular expression of ICAM-1, all of which have been shown in prior in vitro and in vivo murine models to be a direct effect of cocaine metabolic products. Antineutrophilic cytoplasmic antibody and antiphospholipid antibodies are likely the direct sequelae of the proapoptotic microenvironment. The inflammatory vasculitic lesion could reflect the downstream end point reflective of enhanced ICAM-1 expression and the development of antineutrophilic cytoplasmic antibody. Levamisole likely works synergistically with cocaine in the propagation of this syndromic complex. PMID:23392134

  18. Antibodies: an alternative for antibiotics?

    Science.gov (United States)

    Berghman, L R; Abi-Ghanem, D; Waghela, S D; Ricke, S C

    2005-04-01

    In 1967, the success of vaccination programs, combined with the seemingly unstoppable triumph of antibiotics, prompted the US Surgeon General to declare that "it was time to close the books on infectious diseases." We now know that the prediction was overly optimistic and that the fight against infectious diseases is here to stay. During the last 20 yr, infectious diseases have indeed made a staggering comeback for a variety of reasons, including resistance against existing antibiotics. As a consequence, several alternatives to antibiotics are currently being considered or reconsidered. Passive immunization (i.e., the administration of more or less pathogen-specific antibodies to the patient) prior to or after exposure to the disease-causing agent is one of those alternative strategies that was almost entirely abandoned with the introduction of chemical antibiotics but that is now gaining interest again. This review will discuss the early successes and limitations of passive immunization, formerly referred to as "serum therapy," the current use of antibody administration for prophylaxis or treatment of infectious diseases in agriculture, and, finally, recent developments in the field of antibody engineering and "molecular farming" of antibodies in various expression systems. Especially the potential of producing therapeutic antibodies in crops that are routine dietary components of farm animals, such as corn and soy beans, seems to hold promise for future application in the fight against infectious diseases. PMID:15844826

  19. Antibodies to watch in 2016.

    Science.gov (United States)

    Reichert, Janice M

    2016-01-01

    The number of novel antibody therapeutics that received first marketing approvals in 2015 met expectations, with 6 (alirocumab (Praluent®), evolocumab (Repatha®), daratumumab (Darzalex®), dinutuximab (Unituxin®), idarucizumab (Praxbind®), mepolizumab (Nucala®)) granted first approvals as of mid-November*. Seven novel antibody therapeutics (begelomab, brodalumab, elotuzumab, ixekizumab, necitumumab, obiltoxaximab, reslizumab) are in regulatory review, and thus a similar number, if not more, are projected to gain first approvals in 2016. Commercial late-stage antibody therapeutics development exceeded expectations by increasing from 39 candidates in Phase 3 studies as of late 2014 to 53 as of late 2015. Of the 53 candidates, transitions to regulatory review by the end of 2016 are projected for 8 (atezolizumab, benralizumab, bimagrumab, durvalumab, inotuzumab ozogamicin, lebrikizumab, ocrelizumab, tremelimumab). Other "antibodies to watch" include 15 candidates (bavituximab, bococizumab, dupilumab, fasinumab, fulranumab, gevokizumab, guselkumab, ibalizumab, LY2951742, onartuzumab, REGN2222, roledumab, romosozumab, sirukumab, Xilonix) undergoing evaluation in Phase 3 studies that have estimated primary completion dates in 2016. As evidenced by the antibody therapeutics discussed in this perspective, the biopharmaceutical industry has a highly active late-stage clinical pipeline that may deliver numerous new products to the global market in the near future. *See Note added in proof for updates through December 31, 2015. PMID:26651519

  20. The effect of Wolbachia-induced cytoplasmic incompatibility on host population size in natural and manipulated systems.

    OpenAIRE

    Dobson, Stephen L.; Fox, Charles W; Jiggins, Francis M

    2002-01-01

    Obligate, intracellular bacteria of the genus Wolbachia often behave as reproductive parasites by manipulating host reproduction to enhance their vertical transmission. One of these reproductive manipulations, cytoplasmic incompatibility, causes a reduction in egg-hatch rate in crosses between individuals with differing infections. Applied strategies based upon cytoplasmic incompatibility have been proposed for both the suppression and replacement of host populations. As Wolbachia infections ...

  1. Genetic analysis of fertility restoration of Maxie cytoplasmic male sterility im rice (Oryza sativa L. )

    Institute of Scientific and Technical Information of China (English)

    ZHANGXiaoguo; ZHUYingguo; MEIQiming

    1996-01-01

    The genetic behavior of fertility restoration genes of the cytoplasmic male sterile line Maxie A was studied to facilitate the use of Maweizhan male sterile cytoplasm. The F1,F2, and F1 of Maxie A / Minhui 63 were grown in paddy field, 1993-1995.

  2. Characterization of phosphorylation sites in the cytoplasmic domain of the 300 kDa mannose-6-phosphate receptor

    DEFF Research Database (Denmark)

    Rosorius, O; Mieskes, G; Issinger, O G;

    1993-01-01

    kinases phosphorylate the cytoplasmic tail exclusively on serine residues. Inhibition studies using synthetic peptides, partial sequencing of isolated tryptic phosphopeptides and co-migration with tryptic phosphopeptides from MPR 300 labelled in vivo showed that (i) PKA phosphorylates the cytoplasmic MPR...... which may play a role in the transport function of MPR 300 and/or interaction with other proteins....

  3. The molecularly crowded cytoplasm of bacterialcCells : Dividing cells contrasted with viable but non-culturable (VBNC) bacterial cells

    NARCIS (Netherlands)

    Trevors, J. T.; van Elsas, J. D.; Bej, A. K.

    2013-01-01

    In this perspective, we discuss the cytoplasm in actively growing bacterial cells contrasted with viable but non-culturable (VBNC) cells. Actively growing bacterial cells contain a more molecularly crowded and organized cytoplasm, and are capable of completing their cell cycle resulting in cell divi

  4. Inhibition of Insulin Degradation by Hepatoma Cells after Microinjection of Monoclonal Antibodies to a Specific Cytosolic Protease

    Science.gov (United States)

    Shii, Kozui; Roth, Richard A.

    1986-06-01

    Four monoclonal antibodies were identified by their ability to bind to 125I-labeled insulin covalently linked to a cytosolic insulin-degrading enzyme from human erythrocytes. All four antibodies were also found to remove more than 90% of the insulin-degrading activity from erythrocyte extracts. These antibodies were shown to be directed to different sites on the enzyme by mapping studies and by their various properties. Two antibodies recognized the insulin-degrading enzyme from rat liver; one inhibited the erythrocyte enzyme directly; and two recognized the enzyme after gel electrophoresis and transfer to nitrocellulose filters. By this latter procedure and immunoprecipitation from metabolically labeled cells, the enzyme from a variety of tissues was shown to be composed of a single polypeptide chain of apparent Mr 110,000. Finally, these monoclonal antibodies were microinjected into the cytoplasm of a human hepatoma cell line to assess the contribution of this enzyme to insulin degradation in the intact cell. In five separate experiments, preloading of cells with these monoclonal antibodies resulted in an inhibition of insulin degradation of 18-54% (average 39%) and increased the amount of 125I-labeled insulin associated with the cells. In contrast, microinjection of control antibody or an extraneous monoclonal antibody had no effect on insulin degradation or on the amount of insulin associated with the cells. Moreover, the monoclonal antibodies to the insulin-degrading enzyme caused no significant inhibition of degradation of another molecule, low density lipoprotein. Thus, these results support a role for this enzyme in insulin degradation in the intact cell.

  5. Epigenetics of the antibody response.

    Science.gov (United States)

    Li, Guideng; Zan, Hong; Xu, Zhenming; Casali, Paolo

    2013-09-01

    Epigenetic marks, such as DNA methylation, histone post-translational modifications and miRNAs, are induced in B cells by the same stimuli that drive the antibody response. They play major roles in regulating somatic hypermutation (SHM), class switch DNA recombination (CSR), and differentiation to plasma cells or long-lived memory B cells. Histone modifications target the CSR and, possibly, SHM machinery to the immunoglobulin locus; they together with DNA methylation and miRNAs modulate the expression of critical elements of that machinery, such as activation-induced cytidine deaminase (AID), as well as factors central to plasma cell differentiation, such as B lymphocyte-induced maturation protein-1 (Blimp-1). These inducible B cell-intrinsic epigenetic marks instruct the maturation of antibody responses. Their dysregulation plays an important role in aberrant antibody responses to foreign antigens, such as those of microbial pathogens, and self-antigens, such as those targeted in autoimmunity, and B cell neoplasia.

  6. Molecular-specific urokinase antibodies

    Science.gov (United States)

    Atassi, M. Zouhair (Inventor); Morrison, Dennis R. (Inventor)

    2009-01-01

    Antibodies have been developed against the different molecular forms of urokinase using synthetic peptides as immunogens. The peptides were synthesized specifically to represent those regions of the urokinase molecules which are exposed in the three-dimensional configuration of the molecule and are uniquely homologous to urokinase. Antibodies are directed against the lysine 158-isoleucine 159 peptide bond which is cleaved during activation from the single-chain (ScuPA) form to the bioactive double chain (54 KDa and 33 KDa) forms of urokinase and against the lysine 135 lysine 136 bond that is cleaved in the process of removing the alpha-chain from the 54 KDa form to produce the 33 KDa form of urokinase. These antibodies enable the direct measurement of the different molecular forms of urokinase from small samples of conditioned medium harvested from cell cultures.

  7. Tabhu: tools for antibody humanization

    DEFF Research Database (Denmark)

    Olimpieri, Pier Paolo; Marcatili, Paolo; Tramontano, Anna

    2015-01-01

    into a suitable human template. Unfortunately, this procedure may results in a partial or complete loss of affinity of the grafted molecule that can be restored by back-mutating some of the residues of human origin to the corresponding murine ones. This trial-and-error procedure is hard and involves expensive......Antibodies are rapidly becoming essential tools in the clinical practice, given their ability to recognize their cognate antigens with high specificity and affinity, and a high yield at reasonable costs in model animals. Unfortunately, when administered to human patients, xenogeneic antibodies can...... elicit unwanted and dangerous immunogenic responses. Antibody humanization methods are designed to produce molecules with a better safety profile still maintaining their ability to bind the antigen. This can be accomplished by grafting the non-human regions determining the antigen specificity...

  8. Autologous antibodies that bind neuroblastoma cells.

    Science.gov (United States)

    Sun, Yujing; Sholler, Giselle S; Shukla, Girja S; Pero, Stephanie C; Carman, Chelsea L; Zhao, Ping; Krag, David N

    2015-11-01

    Antibody therapy of neuroblastoma is promising and our goal is to derive antibodies from patients with neuroblastoma for developing new therapeutic antibodies. The feasibility of using residual bone marrow obtained for clinical indications as a source of tumor cells and a source of antibodies was assessed. From marrow samples, neuroblastoma cells were recovered, grown in cell culture and also implanted into mice to create xenografts. Mononuclear cells from the marrow were used as a source to generate phage display antibody libraries and also hybridomas. Growth of neuroblastoma patient cells was possible both in vitro and as xenografts. Antibodies from the phage libraries and from the monoclonal hybridomas bound autologous neuroblastoma cells with some selectivity. It appears feasible to recover neuroblastoma cells from residual marrow specimens and to generate human antibodies that bind autologous neuroblastoma cells. Expansion of this approach is underway to collect more specimens, optimize methods to generate antibodies, and to evaluate the bioactivity of neuroblastoma-binding antibodies.

  9. Conformational alterations resulting from mutations in cytoplasmic domains of the alpha subunit of the Na,K-ATPase

    DEFF Research Database (Denmark)

    Blostein, R; Daly, S E; MacAulay, Nanna;

    1998-01-01

    This paper summarizes experiments concerned with the functional consequences of mutations in cytoplasmic regions of the alpha 1 subunit of the Na,K-ATPase, in particular the amino terminus, the first cytoplasmic loop between transmembrane segments M2 and M3, and the major cytoplasmic loop between M...... involving substitution of a residue in the putative cation binding pocket, namely S775A in the fifth transmembrane segment (Arguello, J.M., & Lingrel, J. B. J. Biol. Chem. 270: 22764-22771, 1995). Although its K+/ATP antagonism resembles that of the foregoing cytoplasmic mutants, its vanadate sensitivity is...... unaltered suggesting that changes in apparent affinity for ATP are secondary to changes in K+ ligation. The question of cation selectivity, in particular that of Na+ versus protons, has been addressed in structure/function analysis of a cytoplasmic chimera involving the M4-M5 loop. Transport studies...

  10. Radioimmunotherapy with engineered antibody fragments

    International Nuclear Information System (INIS)

    Authors have developed and begun evaluating radiometal-chelated (213Bi) engineered antibody fragments as radioimmunotherapy agents that target the HER2/neu (c-erbB-2) antigen. The diabody format was found to have 40-fold greater affinity for HER2/neu and to be associated with significantly greater tumor localization than is achieved with scFv molecule. It is shown that short-lived isotopes like 213Bi would be most effective when used in conjunction with antibodies that targeted diffuse malignancies (leukemia or lymphoma) or when used for very rapid pretargeted radioimmunotherapy application in which the radioisotope is conjugated to a very small ligand

  11. Homologous Transcription Factors DUX4 and DUX4c Associate with Cytoplasmic Proteins during Muscle Differentiation.

    Directory of Open Access Journals (Sweden)

    Eugénie Ansseau

    Full Text Available Hundreds of double homeobox (DUX genes map within 3.3-kb repeated elements dispersed in the human genome and encode DNA-binding proteins. Among these, we identified DUX4, a potent transcription factor that causes facioscapulohumeral muscular dystrophy (FSHD. In the present study, we performed yeast two-hybrid screens and protein co-purifications with HaloTag-DUX fusions or GST-DUX4 pull-down to identify protein partners of DUX4, DUX4c (which is identical to DUX4 except for the end of the carboxyl terminal domain and DUX1 (which is limited to the double homeodomain. Unexpectedly, we identified and validated (by co-immunoprecipitation, GST pull-down, co-immunofluorescence and in situ Proximal Ligation Assay the interaction of DUX4, DUX4c and DUX1 with type III intermediate filament protein desmin in the cytoplasm and at the nuclear periphery. Desmin filaments link adjacent sarcomere at the Z-discs, connect them to sarcolemma proteins and interact with mitochondria. These intermediate filament also contact the nuclear lamina and contribute to positioning of the nuclei. Another Z-disc protein, LMCD1 that contains a LIM domain was also validated as a DUX4 partner. The functionality of DUX4 or DUX4c interactions with cytoplasmic proteins is underscored by the cytoplasmic detection of DUX4/DUX4c upon myoblast fusion. In addition, we identified and validated (by co-immunoprecipitation, co-immunofluorescence and in situ Proximal Ligation Assay as DUX4/4c partners several RNA-binding proteins such as C1QBP, SRSF9, RBM3, FUS/TLS and SFPQ that are involved in mRNA splicing and translation. FUS and SFPQ are nuclear proteins, however their cytoplasmic translocation was reported in neuronal cells where they associated with ribonucleoparticles (RNPs. Several other validated or identified DUX4/DUX4c partners are also contained in mRNP granules, and the co-localizations with cytoplasmic DAPI-positive spots is in keeping with such an association. Large muscle RNPs

  12. Secretory TAT-peptide-mediated protein transduction of LIF receptor α-chain distal cytoplasmic motifs into human myeloid HL-60 cells

    Directory of Open Access Journals (Sweden)

    Q. Sun

    2012-10-01

    Full Text Available The distal cytoplasmic motifs of leukemia inhibitory factor receptor α-chain (LIFRα-CT3 can independently induce intracellular myeloid differentiation in acute myeloid leukemia (AML cells by gene transfection; however, there are significant limitations in the potential clinical use of these motifs due to liposome-derived genetic modifications. To produce a potentially therapeutic LIFRα-CT3 with cell-permeable activity, we constructed a eukaryotic expression pcDNA3.0-TAT-CT3-cMyc plasmid with a signal peptide (ss inserted into the N-terminal that codes for an ss-TAT-CT3-cMyc fusion protein. The stable transfection of Chinese hamster ovary (CHO cells via this vector and subsequent selection by Geneticin resulted in cell lines that express and secrete TAT-CT3-cMyc. The spent medium of pcDNA3.0-TAT-CT3-cMyc-transfected CHO cells could be purified using a cMyc-epitope-tag agarose affinity chromatography column and could be detected via SDS-PAGE, with antibodies against cMyc-tag. The direct administration of TAT-CT3-cMyc to HL-60 cell culture media caused the enrichment of CT3-cMyc in the cytoplasm and nucleus within 30 min and led to a significant reduction of viable cells (P < 0.05 8 h after exposure. The advantages of using this mammalian expression system include the ease of generating TAT fusion proteins that are adequately transcripted and the potential for a sustained production of such proteins in vitro for future AML therapy.

  13. Secretory TAT-peptide-mediated protein transduction of LIF receptor α-chain distal cytoplasmic motifs into human myeloid HL-60 cells

    Energy Technology Data Exchange (ETDEWEB)

    Sun, Q. [Department of Hyperbaric Medicine, No. 401 Hospital of PLA, Qingdao (China); Department of Histology and Embryology, Faculty of Basic Medical Sciences, Second Military Medical University, Shanghai (China); Xiong, J. [Department of Histology and Embryology, Faculty of Basic Medical Sciences, Second Military Medical University, Shanghai (China); Lu, J. [Office of Medical Education, Training Department, Second Military Medical University, Shanghai (China); Xu, S. [Department of Histology and Embryology, Faculty of Basic Medical Sciences, Second Military Medical University, Shanghai (China); Li, Y. [State Food and Drug Administration of China,Huangdao Branch, Qingdao (China); Zhong, X.P.; Gao, G.K. [Department of Hyperbaric Medicine, No. 401 Hospital of PLA, Qingdao (China); Liu, H.Q. [2Department of Histology and Embryology, Faculty of Basic Medical Sciences, Second Military Medical University, Shanghai (China)

    2012-06-22

    The distal cytoplasmic motifs of leukemia inhibitory factor receptor α-chain (LIFRα-CT3) can independently induce intracellular myeloid differentiation in acute myeloid leukemia (AML) cells by gene transfection; however, there are significant limitations in the potential clinical use of these motifs due to liposome-derived genetic modifications. To produce a potentially therapeutic LIFRα-CT3 with cell-permeable activity, we constructed a eukaryotic expression pcDNA3.0-TAT-CT3-cMyc plasmid with a signal peptide (ss) inserted into the N-terminal that codes for an ss-TAT-CT3-cMyc fusion protein. The stable transfection of Chinese hamster ovary (CHO) cells via this vector and subsequent selection by Geneticin resulted in cell lines that express and secrete TAT-CT3-cMyc. The spent medium of pcDNA3.0-TAT-CT3-cMyc-transfected CHO cells could be purified using a cMyc-epitope-tag agarose affinity chromatography column and could be detected via SDS-PAGE, with antibodies against cMyc-tag. The direct administration of TAT-CT3-cMyc to HL-60 cell culture media caused the enrichment of CT3-cMyc in the cytoplasm and nucleus within 30 min and led to a significant reduction of viable cells (P < 0.05) 8 h after exposure. The advantages of using this mammalian expression system include the ease of generating TAT fusion proteins that are adequately transcripted and the potential for a sustained production of such proteins in vitro for future AML therapy.

  14. Antibody profiling sensitivity through increased reporter antibody layering

    Energy Technology Data Exchange (ETDEWEB)

    Apel, William A.; Thompson, Vicki S.

    2013-02-26

    A method for analyzing a biological sample by antibody profiling for identifying forensic samples or for detecting the presence of an analyte. In an embodiment of the invention, the analyte is a drug, such as marijuana, Cocaine (crystalline tropane alkaloid), methamphetamine, methyltestosterone, or mesterolone. The method comprises attaching antigens to a surface of a solid support in a preselected pattern to form an array wherein locations of the antigens are known; contacting the array with the biological sample such that a portion of antibodies in the sample reacts with and binds to the antigens in the array to form immune complexes; washing away antibodies that do form immune complexes; and detecting the immune complexes, to form an antibody profile. Forensic samples are identified by comparing a sample from an unknown source with a sample from a known source. Further, an assay, such as a test for illegal drug use, can be coupled to a test for identity such that the results of the assay can be positively correlated to the subject's identity.

  15. Antibody profiling sensitivity through increased reporter antibody layering

    Energy Technology Data Exchange (ETDEWEB)

    Apel, William A; Thompson, Vicki S

    2013-02-26

    A method for analyzing a biological sample by antibody profiling for identifying forensic samples or for detecting the presence of an analyte. In an embodiment of the invention, the analyte is a drug, such as marijuana, Cocaine (crystalline tropane alkaloid), methamphetamine, methyltestosterone, or mesterolone. The method comprises attaching antigens to a surface of a solid support in a preselected pattern to form an array wherein locations of the antigens are known; contacting the array with the biological sample such that a portion of antibodies in the sample reacts with and binds to the antigens in the array to form immune complexes; washing away antibodies that do form immune complexes; and detecting the immune complexes, to form an antibody profile. Forensic samples are identified by comparing a sample from an unknown source with a sample from a known source. Further, an assay, such as a test for illegal drug use, can be coupled to a test for identity such that the results of the assay can be positively correlated to the subject's identity.

  16. Antibody profiling sensitivity through increased reporter antibody layering

    Energy Technology Data Exchange (ETDEWEB)

    Apel, William A.; Thompson, Vicki S

    2010-04-13

    A method for analyzing a biological sample by antibody profiling for identifying forensic samples or for detecting the presence of an analyte. In an embodiment of the invention, the analyte is a drug, such as marijuana, Cocaine (crystalline tropane alkaloid), methamphetamine, methyltestosterone, or mesterolone. The method comprises attaching antigens to a surface of a solid support in a preselected pattern to form an array wherein locations of the antigens are known; contacting the array with the biological sample such that a portion of antibodies in the sample reacts with and binds to the antigens in the array to form immune complexes; washing away antibodies that do form immune complexes; and detecting the immune complexes, to form an antibody profile. Forensic samples are identified by comparing a sample from an unknown source with a sample from a known source. Further, an assay, such as a test for illegal drug use, can be coupled to a test for identity such that the results of the assay can be positively correlated to the subject's identity.

  17. Antibody profiling sensitivity through increased reporter antibody layering

    International Nuclear Information System (INIS)

    A method for analyzing a biological sample by antibody profiling for identifying forensic samples or for detecting the presence of an analyte. In an embodiment of the invention, the analyte is a drug, such as marijuana, Cocaine (crystalline tropane alkaloid), methamphetamine, methyltestosterone, or mesterolone. The method comprises attaching antigens to a surface of a solid support in a preselected pattern to form an array wherein locations of the antigens are known; contacting the array with the biological sample such that a portion of antibodies in the sample reacts with and binds to the antigens in the array to form immune complexes; washing away antibodies that do form immune complexes; and detecting the immune complexes, to form an antibody profile. Forensic samples are identified by comparing a sample from an unknown source with a sample from a known source. Further, an assay, such as a test for illegal drug use, can be coupled to a test for identity such that the results of the assay can be positively correlated to the subject's identity.

  18. Production of biologically active scFv and VHH antibody fragments in Bifidobacterium longum.

    Science.gov (United States)

    Shkoporov, A N; Khokhlova, E V; Savochkin, K A; Kafarskaia, L I; Efimov, B A

    2015-06-01

    Bifidobacteria constitute a significant part of healthy intestinal microbiota in adults and infants and present a promising platform for construction of genetically modified probiotic agents for treatment of gastrointestinal disorders. In this study, three strains of Bifidobacterium longum were constructed that express and secrete biologically active single-chain antibodies against human TNF-α and Clostridium difficile exotoxin A. Anti-TNF-α scFv antibody D2E7 was produced at the level of 25 μg L(-1) in broth culture and was mostly retained in the cytoplasm, while VHH-type antibodies A20.1 and A26.8 against C. difficile exotoxin A were produced at the levels of 0.3-1 mg L(-1) and secreted very efficiently. The biological activity of both antibody types was demonstrated in the mammalian cell-based assays. Expression of A20.1 and A26.8 was also observed in vivo after intragastric administration of transformed B. longum strains to (C57/BL6 × DBA/2)F1 mice. The obtained B. longum strains may serve as prototypes for construction of novel probiotic medications against inflammatory bowel disease and C. difficile-associated disease. PMID:25994292

  19. Migration of antibody secreting cells towards CXCL12 depends on the isotype that forms the BCR

    Science.gov (United States)

    Achatz-Straussberger, Gertrude; Zaborsky, Nadja; Königsberger, Sebastian; Luger, Elke O.; Lamers, Marinus; Crameri, Reto; Achatz, Gernot

    2010-01-01

    Truncation of the cytoplasmic tail of membrane-bound IgE in vivo results in lower serum IgE levels, decreased numbers of IgE-secreting plasma cells and the abrogation of specific secondary immune responses. Here we present mouse strain KN1 that expresses a chimeric ε-γ1 BCR, consisting of the extracellular domains of the ε gene and the trans-membrane and cytoplasmic domains of the γ1 gene. Thus, differences in the IgE immune response of KN1 mice reflect the influence of the “γ1-mediated signalling” of mIgE bearing B cells. KN1 mice show an increased serum IgE level, resulting from an elevated number of IgE-secreting cells. Although the primary IgE immune response in KN1 mice is inconspicuous, the secondary response is far more robust. Most strikingly, IgE-antibody secreting cells with “γ1-signalling history” migrate more efficiently towards the chemokine CXCL12, which guides plasmablasts to plasma cell niches, than IgE-antibody secreting cells with WT “ε-signalling history”. We conclude that IgE plasmablasts have an intrinsic, lower chance to contribute to the long-lived plasma cell pool than IgG1 plasmablasts. PMID:18925577

  20. Ca2+-dependent mobility of vesicles capturing anti-VGLUT1 antibodies

    International Nuclear Information System (INIS)

    Several aspects of secretory vesicle cycle have been studied in the past, but vesicle trafficking in relation to the fusion site is less well understood. In particular, the mobility of recaptured vesicles that traffic back toward the central cytoplasm is still poorly defined. We exposed astrocytes to antibodies against the vesicular glutamate transporter 1 (VGLUT1), a marker of glutamatergic vesicles, to fluorescently label vesicles undergoing Ca2+-dependent exocytosis and examined their number, fluorescence intensity, and mobility by confocal microscopy. In nonstimulated cells, immunolabeling revealed discrete fluorescent puncta, indicating that VGLUT1 vesicles, which are approximately 50 nm in diameter, cycle slowly between the plasma membrane and the cytoplasm. When the cytosolic Ca2+ level was raised with ionomycin, the number and fluorescence intensity of the puncta increased, likely because the VGLUT1 epitopes were more accessible to the extracellularly applied antibodies following Ca2+-triggered exocytosis. In nonstimulated cells, the mobility of labeled vesicles was limited. In stimulated cells, many vesicles exhibited directional mobility that was abolished by cytoskeleton-disrupting agents, indicating dependence on intact cytoskeleton. Our findings show that postfusion vesicle mobility is regulated and may likely play a role in synaptic vesicle cycle, and also more generally in the genesis and removal of endocytic vesicles

  1. Parvovirus particles and movement in the cellular cytoplasm and effects of the cytoskeleton.

    Science.gov (United States)

    Lyi, Sangbom Michael; Tan, Min Jie Alvin; Parrish, Colin R

    2014-05-01

    Cell infection by parvoviruses requires that capsids be delivered from outside the cell to the cytoplasm, followed by genome trafficking to the nucleus. Here we microinject capsids into cells that lack receptors and followed their movements within the cell over time. In general the capsids remained close to the positions where they were injected, and most particles did not move to the vicinity of or enter the nucleus. When 70 kDa-dextran was injected along with the capsids that did not enter the nucleus in significant amounts. Capsids conjugated to peptides containing the SV40 large T-antigen nuclear localization signal remained in the cytoplasm, although bovine serum albumen conjugated to the same peptide entered the nucleus rapidly. No effects of disruption of microfilaments, intermediate filaments, or microtubules on the distribution of the capsids were observed. These results suggest that movement of intact capsids within cells is primarily associated with passive processes.

  2. Parvovirus particles and movement in the cellular cytoplasm and effects of the cytoskeleton

    Energy Technology Data Exchange (ETDEWEB)

    Lyi, Sangbom Michael; Tan, Min Jie Alvin, E-mail: tanmja@gis.a-star.edu.sg; Parrish, Colin R., E-mail: crp3@cornell.edu

    2014-05-15

    Cell infection by parvoviruses requires that capsids be delivered from outside the cell to the cytoplasm, followed by genome trafficking to the nucleus. Here we microinject capsids into cells that lack receptors and followed their movements within the cell over time. In general the capsids remained close to the positions where they were injected, and most particles did not move to the vicinity of or enter the nucleus. When 70 kDa-dextran was injected along with the capsids that did not enter the nucleus in significant amounts. Capsids conjugated to peptides containing the SV40 large T-antigen nuclear localization signal remained in the cytoplasm, although bovine serum albumen conjugated to the same peptide entered the nucleus rapidly. No effects of disruption of microfilaments, intermediate filaments, or microtubules on the distribution of the capsids were observed. These results suggest that movement of intact capsids within cells is primarily associated with passive processes.

  3. A new cytoplasmic male sterile genotype in the sugar beet Beta vulgaris L.: a molecular analysis.

    Science.gov (United States)

    Mann, V; McIntosh, L; Theurer, C; Hirschberg, J

    1989-08-01

    Mitochondrial DNA (mtDNA) from fertile (N) and possibly new cytoplasmic male sterile (CMS) genotypes was studied in the sugar beet Beta vulgaris L. It was found by restriction endonuclease analysis that BMC-CMS, a cytoplasm that was derived from the wild beet Beta maritima, contained a unique type of mtDNA which is distinguishable from both the N and S-CMS, the only other CMS genotype that is currently availabe in B. vulgaris L. The organization of three genes: coxI, coxII and cob, was analyzed by hybridization with heterologous probes from maize. These genes have a similar structure in N and BMC-CMS that is different from S-CMS. It is concluded that BMC-CMS is a novel CMS genotype in the sugar beet.

  4. Actin based processes that could determine the cytoplasmic architecture of plant cells.

    Science.gov (United States)

    van der Honing, Hannie S; Emons, Anne Mie C; Ketelaar, Tijs

    2007-05-01

    Actin polymerisation can generate forces that are necessary for cell movement, such as the propulsion of a class of bacteria, including Listeria, and the protrusion of migrating animal cells. Force generation by the actin cytoskeleton in plant cells has not been studied. One process in plant cells that is likely to depend on actin-based force generation is the organisation of the cytoplasm. We compare the function of actin binding proteins of three well-studied mammalian models that depend on actin-based force generation with the function of their homologues in plants. We predict the possible role of these proteins, and thus the role of actin-based force generation, in the production of cytoplasmic organisation in plant cells.

  5. A cytoplasmic pathway for gapmer antisense oligonucleotide-mediated gene silencing in mammalian cells

    Science.gov (United States)

    Castanotto, Daniela; Lin, Min; Kowolik, Claudia; Wang, LiAnn; Ren, Xiao-Qin; Soifer, Harris S.; Koch, Troels; Hansen, Bo Rode; Oerum, Henrik; Armstrong, Brian; Wang, Zhigang; Bauer, Paul; Rossi, John; Stein, C.A.

    2015-01-01

    Antisense oligonucleotides (ASOs) are known to trigger mRNA degradation in the nucleus via an RNase H-dependent mechanism. We have now identified a putative cytoplasmic mechanism through which ASO gapmers silence their targets when transfected or delivered gymnotically (i.e. in the absence of any transfection reagent). We have shown that the ASO gapmers can interact with the Ago-2 PAZ domain and can localize into GW-182 mRNA-degradation bodies (GW-bodies). The degradation products of the targeted mRNA, however, are not generated by Ago-2-directed cleavage. The apparent identification of a cytoplasmic pathway complements the previously known nuclear activity of ASOs and concurrently suggests that nuclear localization is not an absolute requirement for gene silencing. PMID:26433227

  6. Gravitational effects on the rearrangement of cytoplasmic components during axial formation in amphibian development

    Science.gov (United States)

    Phillips, C. R.; Whalon, B.; Moore, J.; Danilchik, M.

    The spatial positioning of the dorsal-ventral axis in the amphibian, Xenopus laevis, can be experimentally manipulated either by tipping the embryo relative to Earth's gravitational force vector or by centrifugation. Experimental evidence suggests that certain cytoplasmic components are redistributed during the first cell cycle and that these components are, in part, responsible for the establishment of this axis. Further studies indicate that at least some of the cytoplasmic components responsible for establishing this axis may be RNA. Recombinant cDNA and PCR technology are utilized to isolate DNA clones for messenger RNA which becomes spatially localized to the dorsal side of the embryo. These clones are being used to study the mechanisms of spatial localization and the function of the localized RNA transcripts.

  7. Molecular characterization of cytoplasmic male sterility (CMS) in perennial ryegrass ( Lolium perenne L.)

    DEFF Research Database (Denmark)

    Islam, Md. Shofiqul; Møller, Ian Max; Studer, Bruno;

    2011-01-01

    to increase biomass yield, improve nutritional value and tolerance towards abiotic and biotic stress. Cytoplasmic male sterility (CMS) is an efficient tool to control pollination for hybrid seed production. In order to identify the causative polymorphism of the CMS phenotype, a cytoplasmic male sterile plant...... using 454 next-generation sequencing technology, resulting in approximately 800,000 high quality single reads. Here we report on the sequencing and the assembly of the mitochondrial genome from perennial ryegrass. Moreover, the assembly and annotation of the male-sterile and fertile mitochondrial......Perennial ryegrass (Lolium perenne L.) is a key grassland species in temperate climates. As an out breeding crop, ryegrass is currently being bred as population and synthetic families, thereby not fully exploiting the genetically available heterosis. Thus, hybrid breeding has the potential...

  8. Modulation of voltage-gated potassium Kv2.1 via the cytoplasmic C terminal

    Institute of Scientific and Technical Information of China (English)

    Man Jin; Peiyuan Lu

    2011-01-01

    Voltage-gated potassium channels comprise 12 subtypes (Kv1-Kv12). Kv2.1, which is expressed in most mammalian central neurons, provides the majority of delayed-rectifier K current in cortical and hippocampal pyramidal neurons, and plays an especially prominent role in repolarizing membrane potential, as well as in facilitation of exocytosis. Kv2.1-encoded K efflux is essential for neuronal apoptosis programming. The human form of the Kv2.1 potassium channel contains large intracellular regions. The cytoplasmic C-terminal plays a key role in modulating Kv2.1 gating. The present manuscript summarized Kv2.1 structure and modulation in neurons and analyzed the roles of the cytoplasmic C-terminal.

  9. let-65 is cytoplasmic methionyl tRNA synthetase in C. elegans

    Directory of Open Access Journals (Sweden)

    Maha Z. Alriyami

    2014-12-01

    Full Text Available Cytoplasmic methionyl tRNA synthetase (MetRS is one of more than 20 cytoplasmic aminoacyl tRNA synthetase enzymes (ARS. This family of enzymes catalyzes a process fundamental for protein translation. Using a combination of genetic mapping, oligonucleotide array comparative genomic hybridization, and phenotypic correlation, we show that mutations in the essential gene, let-65, reside within the predicted Caenorhabditis elegans homologue of MetRS, which we have named mars-1. We demonstrate that the lethality associated with alleles of let-65 is fully rescued by a transgenic array that spans the mars-1 genomic region. Furthermore, sequence analysis reveals that six let-65 alleles lead to the alteration of highly conserved amino acids.

  10. HER4 and its cytoplasmic isoforms are associated with progression-free survival of malignant melanoma

    DEFF Research Database (Denmark)

    Nielsen, Trine O; Poulsen, Steen Seier; Journe, Fabrice;

    2014-01-01

    RNA quantification was carried out of all four EGF receptors (EGFR, HER2, HER3, and HER4) and the HER4 cytoplasmic isoforms in lymph node metastases from patients with malignant melanoma. We related their expression to progression of the disease. HER4 expression was found to be an indicator of short progression......-free survival (P=0.0340). Interestingly, of the two cytoplasmic splice variants of HER4, the association of CYT1 (P=0.0176) with progression-free survival was more pronounced than that for CYT2 (P=0.0458). Also, HER3 was associated with progression-free survival (P=0.0169), whereas no association was found for...

  11. Pharmacokinetics interactions of monoclonal antibodies.

    Science.gov (United States)

    Ferri, Nicola; Bellosta, Stefano; Baldessin, Ludovico; Boccia, Donatella; Racagni, Giorgi; Corsini, Alberto

    2016-09-01

    The clearance of therapeutic monoclonal antibodies (mAbs) typically does not involve cytochrome P450 (CYP450)-mediated metabolism or interaction with cell membrane transporters, therefore the pharmacokinetics interactions of mAbs and small molecule drugs are limited. However, a drug may affect the clearance of mAbs through the modulation of immune response (e.g., methotrexate reduces the clearance of infliximab, adalimumab, and golimumab, possibly due to methotrexate's inhibitory effect on the formation of antibodies against the mAbs). In addition, mAbs that are cytokine modulators may modify the metabolism of drugs through their effects on P450 enzymes expression. For example, cytokine modulators such as tocilizumab (anti-IL-6 receptor antibody) may reverse the "inhibitory" effect of IL-6 on CYP substrates, resulting in a "normalization" of CYP activities. Finally, a drug may alter the clearance of mAbs by either increasing or reducing the levels of expression of targets of mAbs on the cell surface. For instance, statins and fibrates induce PCSK9 expression and therefore increase cellular uptake and clearance of alirocumab and evolocumab, anti-PCSK9 antibodies. In the present review, we will provide an overview on the pharmacokinetics properties of mAbs as related to the most relevant examples of mAbs-small molecule drug interaction.

  12. The CMS-associated 16 kDa protein encoded by orfH522 in the PET1 cytoplasm is also present in other male-sterile cytoplasms of sunflower.

    Science.gov (United States)

    Horn, R; Hustedt, J E; Horstmeyer, A; Hahnen, J; Zetsche, K; Friedt, W

    1996-02-01

    In sunflower plants carrying the PET1 cytoplasm male sterility (CMS) is associated with a new open reading frame (orfH522) in the 3'-flanking region of the atpA gene and an additional 16 kDa protein. Twenty-seven male-sterile cytoplasms of different origin were studied for the expression of the 16 kDa protein. In addition to the PET1 cytoplasm nine other male-sterile cytoplasms express the CMS-associated protein. These CMS sources originate from different interspecific crosses, from spontaneously occurring male-sterile plants in wild sunflower and from induced mutagenesis. Polyclonal antisera were raised against fusion proteins which contain 421 bp of the 3'-coding region of orfH522 to verify by immunological methods the identity of the other CMS cytoplasms. The anti-ORFH522 antiserum showed a positive reaction in the immunoblot with all CMS cytoplasms which expressing the 16 kDa protein. Investigations of the mitochondrial DNA demonstrated that all ten CMS cytoplasms which express the 16 kDa protein have the same organization at the atpA locus. OrfH522 as probes gave the same transcript pattern for the investigated CMS cytoplasms, just as for PET1. The MAX1 cytoplasm has an orfH522-related sequence but does not synthesize the 16 kDa protein. Using the sodium carbonate treatment the 16 kDa protein proved to be membrane-bound. Computer analyses predict that the hydrophobic N-terminal region of ORFH522 may form a transmembrane helix functioning as membrane anchor. PMID:8605303

  13. Wolbachia-induced cytoplasmic incompatibility as a means for insect pest population control

    OpenAIRE

    Zabalou, Sofia; Riegler, Markus; Theodorakopoulou, Marianna; STAUFFER, CHRISTIAN; Savakis, Charalambos; Bourtzis,Kostas

    2004-01-01

    Biological control is the purposeful introduction of parasites, predators, and pathogens to reduce or suppress pest populations. Wolbachia are inherited bacteria of arthropods that have recently attracted attention for their potential as new biocontrol agents. Wolbachia manipulate host reproduction by using several strategies, one of which is cytoplasmic incompatibility (CI) [Stouthamer, R., Breeuwer, J. A. J. & Hurst, G. D. D. (1999) Annu. Rev. Microbiol. 53, 71–102]. We established Wolbachi...

  14. Isolation and Characterization of the Cytoplasmic Male Sterility Associated Gene of Cotton (Gossypium harknessii)

    Institute of Scientific and Technical Information of China (English)

    WU Jian-yong; GONG Yang-chang; XING Chao-zhu; GUO Li-ping

    2008-01-01

    @@ Cytoplasmic male sterility (CMS) is a maternally inherited trait that results in the failure to produce functional pollen.It was identified in many plants,and it is widely used to exploit heterosis.Now,there are some genes associated with the CMS phenomena that have been identified in the mitochondrial genome.For example,specific genes implicated in CMS have been reported in maize,petunia,bean,Brassica,radish,sunflower,rice,carrot,sorghum,pepper,and many others.

  15. Minimal Plus-end Tracking Unit of the Cytoplasmic Linker Protein CLIP-170*S⃞

    OpenAIRE

    Gupta, Kamlesh K.; Paulson, Benjamin A.; Folker, Eric S.; Charlebois, Blake; Hunt, Alan J.; Goodson, Holly V.

    2009-01-01

    Cytoplasmic linker protein 170 (CLIP-170) is the prototype microtubule (MT) plus-end tracking protein (+TIP) and is involved in regulating MT dynamics. A comprehensive understanding of the process by which CLIP-170 tracks MT plus ends would provide insight into its function. However, the precise molecular mechanism of CLIP-170 +TIP behavior is unknown, and many potential models have been presented. Here, by separating the two CLIP-170 CAP-Gly domains and their adjacent...

  16. Two Stage Cadherin Kinetics Require Multiple Extracellular Domains but Not the Cytoplasmic Region*

    OpenAIRE

    Chien, Yuan-Hung; Jiang, Ning; Li, Fang; Zhang, Fang; Zhu, Cheng; Leckband, Deborah

    2007-01-01

    Micropipette manipulation measurements quantified the pre-steady state binding kinetics between cell pairs mediated by Xenopus cleavage stage cadherin. The time-dependence of the intercellular binding probability exhibits a fast forming, low probability binding state, which transitions to a slower forming, high probability state. The biphasic kinetics are independent of the cytoplasmic region, but the transition to the high probability state requires the third extracellular domain EC3. Deleti...

  17. Intratumoral injection of taxol in vivo suppresses A549 tumor showing cytoplasmic vacuolization.

    Science.gov (United States)

    Wang, Chaoyang; Chen, Tongsheng

    2012-04-01

    Based on our recent in vitro studies, this report was designed to explore the mechanism by which high concentration of taxol (70 µM) induced paraptosis-like cell death in human lung carcinoma (A549) cells, and to evaluate the therapeutic efficacy of taxol using A549 tumor-bearing mice in vivo. Exposure of cells to taxol induced time-dependent cytotoxicity and cytoplasmic vacuolization without the involvement of Bax, Bak, Mcl-1, Bcl-XL, and caspase-3. Although taxol treatment induced activating transcription factor 6 (ATF6) cleavage indicative of endoplasmic reticulum (ER) stress, silencing ATF6 by shATF6 did not prevent taxol-induced both cytotoxcity and cytoplasmic vacuolization, suggesting that taxol-induced cytoplasmic vacuolization and cell death were not due to ER stress. Moreover, taxol-treated cells did not show DNA fragmentation and loss of mitochondrial membrane potential, the typical characteristics of apoptosis. In addition, taxol-induced cytoplasmic vacuolization did not show the cellular lysis, the characteristics of oncosis, and positive of β-galactosidase, the characteristic of senescence, indicating that taxol induced paraptosis-like cell death is neither oncosis nor senescence. Moreover, our in vivo data showed that intratumoral injection of taxol (50 mg/kg) in A549 tumor xenograft mice on day 1 and day 19 potently suppressed tumor growth showing significant ER vacuolization without toxicity. In conclusion, high concentration of taxol exhibits a significant anticancer activity by inducing paraptosis-like cell death in vitro and in vivo, without significant toxicity, suggesting a promising therapeutic strategy for apoptosis-resistance cancer by inducing ER vacuolization.

  18. Relationship between ethylene and the occurrence of cytoplasmic male sterility in rice

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    @@ It was found that exogenous ethylene or ethrel could induce rice male sterility and endogenous ethylene was negatively correlated with pollen fertility and play a key role in the occurrence of rice male sterility. However, it is unknown whether ethylene is related to the occurrence of CMS (cytoplasmic male sterility). In this study, a field experiment was conducted in Guangzhou, using CMS line Zhenshan 97A and its maintainer line Zhenshan 97B.

  19. Actin based processes that could determine the cytoplasmic architecture of plant cells

    OpenAIRE

    Honing; Emons, A.M.C.; Ketelaar, M.J.

    2007-01-01

    Actin polymerisation can generate forces that are necessary for cell movement, such as the propulsion of a class of bacteria, including Listeria, and the protrusion of migrating animal cells. Force generation by the actin cytoskeleton in plant cells has not been studied. One process in plant cells that is likely to depend on actin-based force generation is the organisation of the cytoplasm. We compare the function of actin binding proteins of three well-studied mammalian models that depend on...

  20. Cytoplasmic BRMS1 expression in malignant melanoma is associated with increased disease-free survival

    Directory of Open Access Journals (Sweden)

    Slipicevic Ana

    2012-02-01

    Full Text Available Abstract Background/aims Breast cancer metastasis suppressor 1 (BRMS1 blocks metastasis in melanoma xenografts; however, its usefulness as a biomarker in human melanomas has not been widely studied. The goal was to measure BRMS1 expression in benign nevi, primary and metastatic melanomas and evaluate its impact on disease progression and prognosis. Methods Paraffin-embedded tissue from 155 primary melanomas, 69 metastases and 15 nevi was examined for BRMS1 expression using immunohistochemistry. siRNA mediated BRMS1 down-regulation was used to study impact on invasion and migration in melanoma cell lines. Results A significantly higher percentage of nevi (87%, compared to primary melanomas (20% and metastases (48%, expressed BRMS1 in the nucelus (p Waf1/Cip1 (p = 0.009. Cytoplasmic score index was inversely associated with nuclear p-Akt (p = 0.013 and positively associated with cytoplasmic p-ERK1/2 expression (p = 0.033. Nuclear BRMS1 expression in ≥ 10% of primary melanoma cells was associated with thicker tumors (p = 0.016 and decreased relapse-free period (p = 0.043. Nuclear BRMS1 was associated with expression of fatty acid binding protein 7 (FABP7; p = 0.011, a marker of invasion in melanomas. In line with this, repression of BRMS1 expression reduced the ability of melanoma cells to migrate and invade in vitro. Conclusion Our data suggest that BRMS1 is localized in cytoplasm and nucleus of melanocytic cells and that cellular localization determines its in vivo effect. We hypothesize that cytoplasmic BRMS1 restricts melanoma progression while nuclear BRMS1 possibly promotes melanoma cell invasion. Please see related article: http://www.biomedcentral.com/1741-7015/10/19

  1. Reprogramming of round spermatids by the germinal vesicle cytoplasm in mice.

    Directory of Open Access Journals (Sweden)

    Peng-Cheng Kong

    Full Text Available The birthrate following round spermatid injection (ROSI remains low in current and evidence suggests that factors in the germinal vesicle (GV cytoplasm and certain substances in the GV such as the nucleolus might be responsible for genomic reprogramming and embryonic development. However, little is known whether the reprogramming factors in GV oocyte cytoplasm and/or nucleolus in GV are beneficial to the reprogramming of round spermatids and development of ROSI embryos. Here, round spermatids were treated with GV cytolysates and injected this round spermatid alone or co-injected with GV oocyte nucleolus into mature metaphase II oocytes. Subsequent embryonic development was assessed morphologically and by Oct4 expression in blastocysts. There was no significant difference between experimental groups at the zygote to four-cell development stages. Blastocysts derived from oocytes which were injected with cytolysate treated-round spermatid alone or co-injected with nucleoli injection yielded 63.6% and 70.3% high quality embryos, respectively; comparable to blastocysts derived by intracytoplasmic sperm injection (ICSI, but higher than these oocytes which were co-injected with lysis buffer-treated round spermatids and nucleoli or injected with the lysis buffer-treated round spermatids alone. Furthermore, the proportion of live offspring resulting from oocytes which were co-injected with cytolysate treated-round spermatids and nucleoli or injected with cytolysate treated-round spermatids alone was higher than those were injected with lysis buffer treated-round spermaids, but comparable with the ICSI group. Our results demonstrate that factors from the GV cytoplasm improve round spermatid reprogramming, and while injection of the extra nucleolus does not obviously improve reprogramming its potential contribution, although which cannot be definitively excluded. Thus, some reprogramming factors are evidently present in GV oocyte cytoplasm and could

  2. Vesicles Cytoplasmic Injection: An Efficient Technique to Produce Porcine Transgene-Expressing Embryos.

    Science.gov (United States)

    Luchetti, C G; Bevacqua, R J; Lorenzo, M S; Tello, M F; Willis, M; Buemo, C P; Lombardo, D M; Salamone, D F

    2016-08-01

    The use of vesicles co-incubated with plasmids showed to improve the efficiency of cytoplasmic injection of transgenes in cattle. Here, this technique was tested as a simplified alternative for transgenes delivery in porcine zygotes. To this aim, cytoplasmic injection of the plasmid alone was compared to the injection with plasmids co-incubated with vesicles both in diploid parthenogenic and IVF zygotes. The plasmid pcx-egfp was injected circular (CP) at 3, 30 and 300 ng/μl and linear (LP) at 30 ng/μl. The experimental groups using parthenogenetic zygotes were as follows: CP naked at 3 ng/μl (N = 105), 30 ng/μl (N = 95) and 300 ng/μl (N = 65); Sham (N = 105); control not injected (N = 223); LP naked at 30 ng/μl (N = 78); LP vesicles (N = 115) and Sham vesicles (N = 59). For IVF zygotes: LP naked (N = 44) LP vesicles (N = 94), Sham (N = 59) and control (N = 79). Cleavage, blastocyst and GFP+ rates were analysed by Fisher's test (p plasmids to allow development to blastocyst stage was 30 ng/μl. There were no differences in DNA fragmentation between groups. The parthenogenic LP naked group resulted in high GFP rates (46%) and also allowed the production of GFP blastocysts (33%). The cytoplasmic injection with LP vesicles into parthenogenic zygotes allowed 100% GFP blastocysts. Injected IVF showed higher cleavage rates than control (p GFP blastocysts. The use of vesicles co-incubated with plasmids improves the transgene expression efficiency for cytoplasmic injection in porcine zygotes and constitutes a simple technique for easy delivery of plasmids. PMID:27260090

  3. Why cholesterol should be found predominantly in the cytoplasmic leaf of the plasma membrane

    CERN Document Server

    Giang, Ha

    2014-01-01

    In the mammalian plasma membrane, cholesterol can translocate rapidly between the exoplasmic and cytoplasmic leaves, and is found predominantly in the latter. We hypothesize that it is drawn to the inner leaf to reduce the bending free energy of the membrane caused by the presence there of phosphatidylethanolamine. Incorporating this mechanism into a model free energy for the bilayer, we calculate that approximately two thirds of the total cholesterol should be in the inner leaf.

  4. Diffusion in Cytoplasm: Effects of Excluded Volume Due to Internal Membranes and Cytoskeletal Structures

    OpenAIRE

    Novak, Igor L.; Kraikivski, Pavel; Slepchenko, Boris M.

    2009-01-01

    The intricate geometry of cytoskeletal networks and internal membranes causes the space available for diffusion in cytoplasm to be convoluted, thereby affecting macromolecule diffusivity. We present a first systematic computational study of this effect by approximating intracellular structures as mixtures of random overlapping obstacles of various shapes. Effective diffusion coefficients are computed using a fast homogenization technique. It is found that a simple two-parameter power law prov...

  5. Immunological Interactive Effects between Pollen Grains and Their Cytoplasmic Granules on Brown Norway Rats

    OpenAIRE

    Abou Chakra, Oussama; Jean-Pierre, Sutra; Rogerieux, Françoise; Peltre, Gabriel; Sénéchal, Hélène; Lacroix, Ghislaine

    2009-01-01

    BackgroundGrass pollen is one of the most important aeroallergen vectors in Europe. Under some meteorological factors, pollen grains can release pollen cytoplasmic granules (PCGs). PCGs induce allergic responses. Several studies have shown that during a period of thunderstorms the number of patients with asthma increases because of higher airborne concentrations of PCGs.ObjectiveThe aims of the study were to assess the allergenicity of interactive effects between pollen and PCGs and to compar...

  6. 19 Proteomic Analysis of Major and Minor Allergens From Isolated Pollen Cytoplasmic Granules

    OpenAIRE

    Chakra, Oussama Abou; Sutra, Jean-Pierre; Thomas, Emmanuelle Demey; Vinh, Joëlle; Poncet, Pascal; Lacroix, Ghislaine; Sénéchal, Hélène

    2012-01-01

    Background Grass pollen is one of the most important vectors of aeroallergens. Under atmospheric conditions, pollen grains can release pollen cytoplasmic granules (PCGs). The allergens associated with these intrinsic sub-fractions induce, in laboratory animals as well as in asthmatic patients allergic and inflammatory responses. The aims of this study were to characterize and identify the intrinsic allergens of PCGs, to compare them with those of pollen grains. Methods PCGs were isolated from...

  7. Crystal Structure of the Pseudomonas aeruginosa Cytoplasmic Heme Binding Protein, Apo-PhuS

    OpenAIRE

    Tripathi, Sarvind; O'Neill, Maura J.; Wilks, Angela; Poulos, Thomas L.

    2013-01-01

    Iron is an essential element to all living organisms and is an important determinant of bacterial virulence. Bacteria have evolved specialized systems to sequester and transport iron from the environment or host. Pseudomonas aeruginosa, an opportunistic pathogen, uses two outer membrane receptor mediated systems (Phu and Has) to utilize host heme as a source of iron. PhuS is a 39 kDa soluble cytoplasmic heme binding protein which interacts and transports heme from the inner membrane heme tran...

  8. Cytoplasmic male sterility contributes to hybrid incompatibility between subspecies of Arabidopsis lyrata.

    Science.gov (United States)

    Aalto, Esa A; Koelewijn, Hans-Peter; Savolainen, Outi

    2013-10-03

    In crosses between evolutionarily diverged populations, genomic incompatibilities may result in sterile hybrids, indicating evolution of reproductive isolation. In several plant families, crosses within a population can also lead to male sterile progeny because of conflict between the maternally and biparentally inherited genomes. We examined hybrid fertility between subspecies of the perennial outcrossing self-incompatible Lyrate rockcress (Arabidopsis lyrata) in large reciprocal F2 progenies and three generations of backcrosses. In one of the reciprocal F2 progenies, almost one-fourth of the plants were male-sterile. Correspondingly, almost one-half of the plants in one of the four reciprocal backcross progenies expressed male sterility. In an additional four independent F2 and backcross families, three segregated male sterility. The observed asymmetrical hybrid incompatibility is attributable to male sterility factors in one cytoplasm, for which the other population lacks effective fertility restorers. Genotyping of 96 molecular markers and quantitative trait locus mapping revealed that only 60% of the plants having the male sterile cytoplasm and lacking the corresponding restorers were phenotypically male-sterile. Genotyping data showed that there is only one restorer locus, which mapped to a 600-kb interval at the top of chromosome 2 in a region containing a cluster of pentatricopeptide repeat genes. Male fertility showed no trade-off with seed production. We discuss the role of cytoplasm and genomic conflict in incipient speciation and conclude that cytoplasmic male sterility-lowering hybrid fitness is a transient effect with limited potential to form permanent reproductive barriers between diverged populations of hermaphrodite self-incompatible species.

  9. The Architecture of the Cytoplasmic Region of Type III Secretion Systems

    Science.gov (United States)

    Makino, Fumiaki; Shen, Dakang; Kajimura, Naoko; Kawamoto, Akihiro; Pissaridou, Panayiota; Oswin, Henry; Pain, Maria; Murillo, Isabel; Namba, Keiichi; Blocker, Ariel J.

    2016-01-01

    Type III secretion systems (T3SSs) are essential devices in the virulence of many Gram-negative bacterial pathogens. They mediate injection of protein effectors of virulence from bacteria into eukaryotic host cells to manipulate them during infection. T3SSs involved in virulence (vT3SSs) are evolutionarily related to bacterial flagellar protein export apparatuses (fT3SSs), which are essential for flagellar assembly and cell motility. The structure of the external and transmembrane parts of both fT3SS and vT3SS is increasingly well-defined. However, the arrangement of their cytoplasmic and inner membrane export apparatuses is much less clear. Here we compare the architecture of the cytoplasmic regions of the vT3SSs of Shigella flexneri and the vT3SS and fT3SS of Salmonella enterica serovar Typhimurium at ~5 and ~4 nm resolution using electron cryotomography and subtomogram averaging. We show that the cytoplasmic regions of vT3SSs display conserved six-fold symmetric features including pods, linkers and an ATPase complex, while fT3SSs probably only display six-fold symmetry in their ATPase region. We also identify other morphological differences between vT3SSs and fT3SSs, such as relative disposition of their inner membrane-attached export platform, C-ring/pods and ATPase complex. Finally, using classification, we find that both types of apparatuses can loose elements of their cytoplasmic region, which may therefore be dynamic. PMID:27686865

  10. Cytoplasmic pH and the regulation of the dictyostelium cell cycle

    NARCIS (Netherlands)

    Aerts, R.J.; Durston, A.J.; Moolenaar, W.H.

    1985-01-01

    Cytoplasmic pH (pHl) was monitored during the cell cycle of synchronous populations of Dictyostelium discoideum by means of a pH “null point” method. There is a cycle of pHl that closely corresponds to the DNA replication cycle, with a minimum of pH 7.20 in interphase and a peak of pH 7.45 during S

  11. Radiation Hybrid Mapping of the Species Cytoplasm-Specific (scsae) Gene in Wheat

    OpenAIRE

    Hossain, Khwaja G.; Riera-Lizarazu, Oscar; Kalavacharla, Venugopal; Vales, M. Isabel; Maan, Schivcharan S.; Kianian, Shahryar F

    2004-01-01

    Radiation hybrid (RH) mapping is based on radiation-induced chromosome breakage and analysis of chromosome segment retention or loss using molecular markers. In durum wheat (Triticum turgidum L., AABB), an alloplasmic durum line [(lo) durum] has been identified with chromosome 1D of T. aestivum L. (AABBDD) carrying the species cytoplasm-specific (scsae) gene. The chromosome 1D of this line segregates as a whole without recombination, precluding the use of conventional genome mapping. A radiat...

  12. Aldolase exists in both the fluid and solid phases of cytoplasm.

    Science.gov (United States)

    Pagliaro, L; Taylor, D L

    1988-09-01

    We have prepared a functional fluorescent analogue of the glycolytic enzyme aldolase (rhodamine [Rh]-aldolase), using the succinimidyl ester of carboxytetramethyl-rhodamine. Fluorescence redistribution after photobleaching measurements of the diffusion coefficient of Rh-aldolase in aqueous solutions gave a value of 4.7 x 10(-7) cm2/S, and no immobile fraction. In the presence of filamentous actin, there was a 4.5-fold reduction in diffusion coefficient, as well as a 36% immobile fraction, demonstrating binding of Rh-aldolase to actin. However, in the presence of a 100-fold molar excess of its substrate, fructose 1,6-diphosphate, both the mobile fraction and diffusion coefficient of Rh-aldolase returned to control levels, indicating competition between substrate binding and actin cross-linking. When Rh-aldolase was microinjected into Swiss 3T3 cells, a relatively uniform intracellular distribution of fluorescence was observed. However, there were significant spatial differences in the in vivo diffusion coefficient and mobile fraction of Rh-aldolase measured with fluorescence redistribution after photobleaching. In the perinuclear region, we measured an apparent cytoplasmic diffusion coefficient of 1.1 x 10(-7) cm2/s with a 23% immobile fraction; while measurements in the cell periphery gave a value of 5.7 x 10(-8) cm2/s, with no immobile fraction. Ratio imaging of Rh-aldolase and FITC-dextran indicated that FITC-dextran was relatively excluded excluded from stress fiber domains. We interpret these data as evidence for the partitioning of aldolase between a soluble fraction in the fluid phase and a fraction associated with the solid phase of cytoplasm. The partitioning of aldolase and other glycolytic enzymes between the fluid and solid phases of cytoplasm could play a fundamental role in the control of glycolysis, the organization of cytoplasm, and cell motility. The concepts and experimental approaches described in this study can be applied to other cellular

  13. Self-fertilization and restoration to cytoplasmic male sterility of some wild species of helianthus

    OpenAIRE

    Onemli F.; Gucer T.

    2010-01-01

    The objectives of this study were to determine self-fertility and restoration capability of cytoplasmic male fertility of some wild sunflower genotypes. The evaluated genotypes were Helianthus petiolaris spp. petiolaris (E-142), Helianthus neglectus (E-017) and Helianthus annuus (E-060, E-173, E-174 and E-175). For hybridization, the female inbred line 2453-A was used. Results showed that self fertility ratios of wild sunflower genotypes varied between 22.4...

  14. The Active Site of Oligogalacturonate Lyase Provides Unique Insights into Cytoplasmic Oligogalacturonate β-Elimination*

    OpenAIRE

    Abbott, D. Wade; Gilbert, Harry J.; Boraston, Alisdair B.

    2010-01-01

    Oligogalacturonate lyases (OGLs; now also classified as pectate lyase family 22) are cytoplasmic enzymes found in pectinolytic members of Enterobacteriaceae, such as the enteropathogen Yersinia enterocolitica. OGLs utilize a β-elimination mechanism to preferentially catalyze the conversion of saturated and unsaturated digalacturonate into monogalacturonate and the 4,5-unsaturated monogalacturonate-like molecule, 5-keto-4-deoxyuronate. To provide mechanistic insights into the specificity of th...

  15. Development of a New Excellent Cytoplasmic Type of CMS Line in Rice

    Institute of Scientific and Technical Information of China (English)

    FAN Shu-guo; LIANG Cheng-ye; LIU Hong-xian

    2001-01-01

    A new excellent cytoplasmic type of CMS line Sl0-2A in rice with typical abortion type (TAT)from Minghui 63 was obtained by in vitro culture. It possesses high percentage of exerted stigma with strong vitality, big opening angle of spikelet, high flowering rate before noon and iow percentage of cleistogamous spikelet. The relationship between restoring and maintaining of S10-2A is completely different from that of WA type, HL type and BT type.

  16. Differential effect of HDAC3 on cytoplasmic and nuclear huntingtin aggregates.

    Directory of Open Access Journals (Sweden)

    Tatsuo Mano

    Full Text Available Histone deacetylases (HDACs are potential therapeutic targets of polyglutamine (pQ diseases including Huntington's disease (HD that may function to correct aberrant transcriptional deactivation caused by mutant pQ proteins. HDAC3 is a unique class 1 HDAC found in both the cytoplasm and in the nucleus. However, the precise functions of HDAC3 in the two cellular compartments are only vaguely known. HDAC3 directly binds to huntingtin (Htt with short pQ and this interaction is important for suppressing neurotoxicity induced by HDAC3. With long pQ Htt, the interaction with HDAC3 is inhibited, and this supposedly promotes neuronal death, indicating that HDAC3 would be a good therapeutic target for HD. However, the knockout of one HDAC3 allele did not show any efficacy in reducing neurodegenerative symptoms in a mouse model of HD. Therefore, the role of HDAC3 in the pathogenesis of HD has yet to be fully elucidated. We attempted to resolve this issue by focusing on the different roles of HDAC3 on cytoplasmic and nuclear Htt aggregates. In addition to supporting the previous findings, we found that HDAC3 preferentially binds to nuclear Htt over cytoplasmic ones. Specific HDAC3 inhibitors increased the total amount of Htt aggregates by increasing the amount of nuclear aggregates. Both cytoplasmic and nuclear Htt aggregates were able to suppress endogenous HDAC3 activity, which led to decreased nuclear proteasome activity. Therefore, we concluded that Htt aggregates impair nuclear proteasome activity through the inhibition of HDAC3. Our findings provide new insights regarding cross-compartment proteasome regulation.

  17. The role of cytoplasmic catalase in dehydration tolerance of Saccharomyces cerevisiae

    OpenAIRE

    França, Mauro Braga; Panek, Anita Dolly; Eleutherio, Elis Cristina Araujo

    2005-01-01

    In this study, we investigated the role played by cytoplasmic catalase (Ctt1) in resistance against water loss using the yeast Saccharomyces cerevisiae as eukaryotic cell model. Comparing a mutant possessing a specific lesion in CTT1 with its parental strain, it was observed that both control and ctt1 strains exhibited increased levels of lipid peroxidation after dehydration, suggesting that catalase does not protect membranes during drying. Although the ctt1 strain has only 1 catalase isofor...

  18. Generation of micronuclei during interphase by coupling between cytoplasmic membrane blebbing and nuclear budding.

    Directory of Open Access Journals (Sweden)

    Koh-ichi Utani

    Full Text Available Micronucleation, mediated by interphase nuclear budding, has been repeatedly suggested, but the process is still enigmatic. In the present study, we confirmed the previous observation that there are lamin B1-negative micronuclei in addition to the positive ones. A large cytoplasmic bleb was found to frequently entrap lamin B1-negative micronuclei, which were connected to the nucleus by a thin chromatin stalk. At the bottom of the stalk, the nuclear lamin B1 structure appeared broken. Chromatin extrusion through lamina breaks has been referred to as herniation or a blister of the nucleus, and has been observed after the expression of viral proteins. A cell line in which extrachromosomal double minutes and lamin B1 protein were simultaneously visualized in different colors in live cells was established. By using these cells, time-lapse microscopy revealed that cytoplasmic membrane blebbing occurred simultaneously with the extrusion of nuclear content, which generated lamin B1-negative micronuclei during interphase. Furthermore, activation of cytoplasmic membrane blebbing by the addition of fresh serum or camptothecin induced nuclear budding within 1 to 10 minutes, which suggested that blebbing might be the cause of the budding. After the induction of blebbing, the frequency of lamin-negative micronuclei increased. The budding was most frequent during S phase and more efficiently entrapped small extrachromosomal chromatin than the large chromosome arm. Based on these results, we suggest a novel mechanism in which cytoplasmic membrane dynamics pulls the chromatin out of the nucleus through the lamina break. Evidence for such a mechanism was obtained in certain cancer cell lines including human COLO 320 and HeLa. The mechanism could significantly perturb the genome and influence cancer cell phenotypes.

  19. Plant cytoplasmic GAPDH: redox post-translational modifications and moonlighting properties

    Directory of Open Access Journals (Sweden)

    Mirko eZaffagnini

    2013-11-01

    Full Text Available Glyceraldehyde-3-phosphate dehydrogenase (GAPDH is a ubiquitous enzyme involved in glycolysis and shown, particularly in animal cells, to play additional roles in several unrelated non-metabolic processes such as control of gene expression and apoptosis. This functional versatility is regulated, in part at least, by redox post-translational modifications that alter GAPDH catalytic activity and influence the subcellular localization of the enzyme. In spite of the well established moonlighting (multifunctional properties of animal GAPDH, little is known about non-metabolic roles of GAPDH in plants. Plant cells contain several GAPDH isoforms with different catalytic and regulatory properties, located both in the cytoplasm and in plastids, and participating in glycolysis and the Calvin-Benson cycle. A general feature of all GAPDH proteins is the presence of an acidic catalytic cysteine in the active site that is overly sensitive to oxidative modifications, including glutathionylation and S-nitrosylation. In Arabidopsis, oxidatively-modified cytoplasmic GAPDH has been successfully used as a tool to investigate the role of reduced glutathione, thioredoxins and glutaredoxins in the control of different types of redox post-translational modifications. Oxidative modifications inhibit GAPDH activity, but might enable additional functions in plant cells. Mounting evidence support the concept that plant cytoplasmic GAPDH may fulfill alternative, non-metabolic functions that are triggered by redox post-translational modifications of the protein under stress conditions. The aim of this review is to detail the molecular mechanisms underlying the redox regulation of plant cytoplasmic GAPDH in the light of its crystal structure, and to provide a brief inventory of the well known redox-dependent multi-facetted properties of animal GAPDH, together with the emerging roles of oxidatively-modified GAPDH in stress signaling pathways in plants.

  20. HDAC4 reduction: a novel therapeutic strategy to target cytoplasmic huntingtin and ameliorate neurodegeneration.

    Directory of Open Access Journals (Sweden)

    Michal Mielcarek

    2013-11-01

    Full Text Available Histone deacetylase (HDAC 4 is a transcriptional repressor that contains a glutamine-rich domain. We hypothesised that it may be involved in the molecular pathogenesis of Huntington's disease (HD, a protein-folding neurodegenerative disorder caused by an aggregation-prone polyglutamine expansion in the huntingtin protein. We found that HDAC4 associates with huntingtin in a polyglutamine-length-dependent manner and co-localises with cytoplasmic inclusions. We show that HDAC4 reduction delayed cytoplasmic aggregate formation, restored Bdnf transcript levels, and rescued neuronal and cortico-striatal synaptic function in HD mouse models. This was accompanied by an improvement in motor coordination, neurological phenotypes, and increased lifespan. Surprisingly, HDAC4 reduction had no effect on global transcriptional dysfunction and did not modulate nuclear huntingtin aggregation. Our results define a crucial role for the cytoplasmic aggregation process in the molecular pathology of HD. HDAC4 reduction presents a novel strategy for targeting huntingtin aggregation, which may be amenable to small-molecule therapeutics.

  1. A detrimental mitochondrial-nuclear interaction causes cytoplasmic male sterility in rice.

    Science.gov (United States)

    Luo, Dangping; Xu, Hong; Liu, Zhenlan; Guo, Jingxin; Li, Heying; Chen, Letian; Fang, Ce; Zhang, Qunyu; Bai, Mei; Yao, Nan; Wu, Hong; Wu, Hao; Ji, Chonghui; Zheng, Huiqi; Chen, Yuanling; Ye, Shan; Li, Xiaoyu; Zhao, Xiucai; Li, Riqing; Liu, Yao-Guang

    2013-05-01

    Plant cytoplasmic male sterility (CMS) results from incompatibilities between the organellar and nuclear genomes and prevents self pollination, enabling hybrid crop breeding to increase yields. The Wild Abortive CMS (CMS-WA) has been exploited in the majority of 'three-line' hybrid rice production since the 1970s, but the molecular basis of this trait remains unknown. Here we report that a new mitochondrial gene, WA352, which originated recently in wild rice, confers CMS-WA because the protein it encodes interacts with the nuclear-encoded mitochondrial protein COX11. In CMS-WA lines, WA352 accumulates preferentially in the anther tapetum, thereby inhibiting COX11 function in peroxide metabolism and triggering premature tapetal programmed cell death and consequent pollen abortion. WA352-induced sterility can be suppressed by two restorer-of-fertility (Rf) genes, suggesting the existence of different mechanisms to counteract deleterious cytoplasmic factors. Thus, CMS-related cytoplasmic-nuclear incompatibility is driven by a detrimental interaction between a newly evolved mitochondrial gene and a conserved, essential nuclear gene.

  2. Region of Nipah virus C protein responsible for shuttling between the cytoplasm and nucleus.

    Science.gov (United States)

    Horie, Ryo; Yoneda, Misako; Uchida, Shotaro; Sato, Hiroki; Kai, Chieko

    2016-10-01

    Nipah virus (NiV) causes severe encephalitis in humans, with high mortality. NiV nonstructural C protein (NiV-C) is essential for its pathogenicity, but its functions are unclear. In this study, we focused on NiV-C trafficking in cells and found that it localizes predominantly in the cytoplasm but partly in the nucleus. An analysis of NiV-C mutants showed that amino acids 2, 21-24 and 110-139 of NiV-C are important for its localization in the cytoplasm. Inhibitor treatment indicates that the nuclear export determinant is not a classical CRM1-dependent nuclear export signal. We also determined that amino acids 60-75 and 72-75 were important for nuclear localization of NiV-C. Furthermore, NiV-C mutants that had lost their capacity for nuclear localization inhibited the interferon (IFN) response more strongly than complete NiV-C. These results indicate that the IFN-antagonist activity of NiV-C occurs in the cytoplasm.

  3. Cooperative regulation of cytoplasmic streaming and ca fluxes by pfr and photosynthesis in vallisneria mesophyll cells.

    Science.gov (United States)

    Takagi, S; Yamamoto, K T; Furuya, M; Nagai, R

    1990-12-01

    In mesophyll cells of Vallisneria gigantea Graebner, Ca(2+) regulates the induction and cessation of cytoplasmic streaming. Streaming is induced when the level of calcium in the cytoplasm is lowered through light-accelerated release of Ca(2+) from the cells (S Takagi, R Nagai [1988] Plant Physiol 88: 228-232). We have now initiated an investigation on the nature of the photoreceptor(s) that are involved in the regulation of Ca(2+) movements across the cell membrane and of streaming. Streaming is induced only when phytochrome exists in the phytochrome-far redabsorbing form (Pfr)-and photosynthesis is allowed to take place for at least 4 minutes. The former effect is typically photoreversible by red and far-red light, and phytochrome is spectro-photometrically detectable in the crude extract from the leaves. The latter effect is assessed in terms of the wavelength dependency and the effects of diuron and atrazine, two inhibitors of photosynthesis. A similar requirement for Pfr and photosynthesis is found to be associated with the acceleration of Ca(2+) efflux in the protoplasts. The results suggest that phytochrome and photosynthetic pigment(s) cooperatively regulate cytoplasmic streaming via modulation of the Ca(2+) transport in the cell membrane. PMID:16667905

  4. Characterization of Nuclear and Cytoplasmic Compositions of Somatic Hybrid Plants Between Sweet Orange and Sour Orange

    Institute of Scientific and Technical Information of China (English)

    LIUJi-Hong; XUXiao-Yong; DENGXiu-Xin

    2004-01-01

    Abstract In the present research, flow cytometry (FCM), simple sequence repeat (SSR) and cleavedamplified polymorphic sequence (CAPS) were employed to analyze somatic hybrid plants derived from electrofusion between embryogenic protoplasts of sweet orange (Citrus sinensis Osbeck cv. Shamouti) and leaf-derived protoplasts of sour orange (C. aurantium L). FCM showed that all of the somatic hybrid plants had two-fold fluorescence intensity values of the diploid control, indicating that they were tetraploids. SSR and CAPS were used to characterize the compositions of nuclear and cytoplasmic genomes of the somatic hybnd plants. As for SSR four primer pairs were tried and two showed polymorphisms between the fusion parents. With both pnmer pairs the somatic hybrid plants encompassed DNA bands from both parents, indicating that they were heterokaryonic hybrids. Amplification with some universal primers, followed by digestion with restriction endonucleases, could distinguish the fusion parents from each other. As far as mitochondrial and chloroplast DNA compositions were concerned the somatic hybrid plants shared the same banding patterns as the embryogenic parents for all of the polymorphic primer/enzyme combinations. The results herein demonstrated that the somatic hybrid plants inherited their nuclear genome from both fusion parents, whereas the cytoplasmic genomes were possibly only contributed by the embryogenic parent. Merits of the analytical methods and nuclear and cytoplasmic inheritance of citrus tetraploid somatic hybrids, together with their application, are discussed herein.

  5. The cytoplasmic extension of the integrin β6 subunit regulates epithelial-to-mesenchymal transition.

    Science.gov (United States)

    Lee, Carlin; Lee, Casey; Lee, Stacey; Siu, Amanda; Ramos, Daniel M

    2014-02-01

    Prognosis for oral cancer patients has not improved in over 60 years due to invasion and recurrence. To understand the invasive behavior of this tumor, we evaluated the role of the αvβ6 integrin. Invasive oral SCC cells express the αvβ6 integrin, which contains an 11-amino-acid extension on its β-subunit unique to the integrin family. We determined that this β6 cytoplasmic extension regulates the composition of the intermediate filament network and the organization of signaling structures called focal contacts. The auto-phosphorylation of FAK, which is localized to focal contacts, was also regulated by the β6-cytoplasmic tail, as were the transcription factors Notch and STAT3. Lastly, we also determined that activation of MAPK required the full-length β6 integrin. Together these results indicate that the signaling critical to epithelial-to-mesenchymal transition (EMT) is regulated by the β6 integrin cytoplasmic domain. PMID:24510996

  6. Alternative affinity tools: more attractive than antibodies?

    NARCIS (Netherlands)

    Ruigrok, V.J.B.; Levisson, M.; Eppink, M.H.M.; Smidt, H.; Oost, van der J.

    2011-01-01

    Antibodies are the most successful affinity tools used today, in both fundamental and applied research (diagnostics, purification and therapeutics). Nonetheless, antibodies do have their limitations, including high production costs and low stability. Alternative affinity tools based on nucleic acids

  7. Detection of Campylobacter species using monoclonal antibodies

    Science.gov (United States)

    Young, Colin R.; Lee, Alice; Stanker, Larry H.

    1999-01-01

    A panel of species specific monoclonal antibodies were raised to Campylobacter coli, Campylobacter jejuni and Campylobacter lari. The isotypes, and cross-reactivity profiles of each monoclonal antibody against an extensive panel of micro- organisms, were determined.

  8. [Neuroimmunological diseases associated with VGKC complex antibodies].

    Science.gov (United States)

    Watanabe, Osamu

    2013-05-01

    Antibodies to voltage-gated potassium channels(VGKC) were first identified by radioimmunoassay of radioisotope labeled alpha-dendrotoxin-VGKCs solubilized from rabbit brain. These antibodies were found only in a proportion of patients with acquired neuromyotonia (Isaacs' syndrome). VGKC antibodies were also detected in Morvan's syndrome and in a form of autoimmune limbic encephalitis. Recent studies indicated that the "VGKC" antibodies are mainly directed toward associated proteins(for example LGI-1, Caspr-2) that complex with the VGKCs themselves. The "VGKC" antibodies are now usually known as VGKC-complex antibodies. In general, LGI-1 antibodies are most common in limbic encephalitis with SIADH. Caspr-2 antibodies are present in the majority of patients with Morvan's syndrome. These patients develop combinations of CNS symptoms, autonomic dysfunction, and peripheral nerve hyperexcitability.

  9. A new source of cytoplasmic male sterility found in wild beet and its relationship to other CMS types.

    Science.gov (United States)

    Kawanishi, Yuki; Shinada, Hiroshi; Matsunaga, Muneyuki; Masaki, Yusuke; Mikami, Tetsuo; Kubo, Tomohiko

    2010-04-01

    We found a number of male-sterile plants in a wild beet (Beta vulgaris L. subsp. maritima) accession line, FR4-31. The inheritance study of the male sterility indicated the trait to be of the cytoplasmic type. The mitochondrial genome of FR4-31 proved to lack the male-sterility-associated genes preSatp6 and orf129, which are characteristic of the Owen CMS and I-12CMS(3) cytoplasms of beets, respectively. Instead, the truncated cox2 gene involved in G CMS originating from wild beets was present in the FR4-31 mitochondrial genome. In Southern hybridization using four mitochondrial gene probes, the FR4-31 cytoplasm showed patterns similar to those typical of the G cytoplasm. It is thus likely that the FR4-31 cytoplasm has a different CMS mechanism from both Owen CMS and I-12CMS(3), and that the FR4-31 and G cytoplasms resemble each other closely. A restriction map of the FR4-31 mitochondrial DNA was generated and aligned with those published for the Owen and normal fertile cytoplasms. The FR4-31 mitochondrial genome was revealed to differ extensively in arrangement from the Owen and normal genomes, and the male-sterile Owen and FR4-31 genomes seem to be derived independently from an ancestral genome.

  10. A new source of cytoplasmic male sterility found in wild beet and its relationship to other CMS types.

    Science.gov (United States)

    Kawanishi, Yuki; Shinada, Hiroshi; Matsunaga, Muneyuki; Masaki, Yusuke; Mikami, Tetsuo; Kubo, Tomohiko

    2010-04-01

    We found a number of male-sterile plants in a wild beet (Beta vulgaris L. subsp. maritima) accession line, FR4-31. The inheritance study of the male sterility indicated the trait to be of the cytoplasmic type. The mitochondrial genome of FR4-31 proved to lack the male-sterility-associated genes preSatp6 and orf129, which are characteristic of the Owen CMS and I-12CMS(3) cytoplasms of beets, respectively. Instead, the truncated cox2 gene involved in G CMS originating from wild beets was present in the FR4-31 mitochondrial genome. In Southern hybridization using four mitochondrial gene probes, the FR4-31 cytoplasm showed patterns similar to those typical of the G cytoplasm. It is thus likely that the FR4-31 cytoplasm has a different CMS mechanism from both Owen CMS and I-12CMS(3), and that the FR4-31 and G cytoplasms resemble each other closely. A restriction map of the FR4-31 mitochondrial DNA was generated and aligned with those published for the Owen and normal fertile cytoplasms. The FR4-31 mitochondrial genome was revealed to differ extensively in arrangement from the Owen and normal genomes, and the male-sterile Owen and FR4-31 genomes seem to be derived independently from an ancestral genome. PMID:20616856

  11. The cyclin D1 proto-oncogene is sequestered in the cytoplasm of mammalian cancer cell lines

    Directory of Open Access Journals (Sweden)

    Coombes R Charles

    2006-02-01

    Full Text Available Abstract Background The cyclin D1 proto-oncogene is an important regulator of G1 to S-phase transition and an important cofactor for several transcription factors in numerous cell types. Studies on neonatal cardiomyocytes and postmitotic neurons indicate that the activity of cyclin D1 may be regulated through its cytoplasmic sequestration. We have demonstrated previously, that TSA induces the ubiquitin-dependent degradation of cyclin D1 in MCF-7 breast cancer cells. Additional studies were initiated in order to further investigate the effect of TSA on cyclin D1 regulation using sub-cellular fractionation techniques. Results Our studies revealed cyclin D1 to be localized predominantly within the cytoplasmic fraction of all cell lines tested. These observations were confirmed by confocal microscopy. GSK3β was found to be localized within both the nucleus and cytoplasm throughout the cell cycle. Inhibition of GSK3β or CRM1-dependent nuclear export resulted in only modest nuclear accumulation, suggesting that the cytoplasmic localization of cyclin D1 results from the inhibition of its nuclear import. Conclusion We have shown by several different experimental approaches, that cyclin D1 is in fact a predominantly cytoplasmic protein in mammalian cancer cell lines. Recent studies have shown that the cytoplasmic sequestration of cyclin D1 prevents apoptosis in neuronal cells. Our results suggest that cytoplasmic sequestration may additionally serve to regulate cyclin D1 activity in mammalian cancer cells.

  12. Classification of Cells with Membrane Staining and/or Fixation Based on Cellular Specific Membrane Capacitance and Cytoplasm Conductivity

    Directory of Open Access Journals (Sweden)

    Song-Bin Huang

    2015-01-01

    Full Text Available Single-cell electrical properties (e.g., specific membrane capacitance (Cspecific membrane and cytoplasm conductivity (σcytoplasm have been regarded as potential label-free biophysical markers for the evaluation of cellular status. However, whether there exist correlations between these biophysical markers and cellular status (e.g., membrane-associate protein expression is still unknown. To further validate the utility of single-cell electrical properties in cell type classification, Cspecific membrane and σcytoplasm of single PC-3 cells with membrane staining and/or fixation were analyzed and compared in this study. Four subtypes of PC-3 cells were prepared: untreated PC-3 cells, PC-3 cells with anti-EpCAM staining, PC-3 cells with fixation, and fixed PC-3 cells with anti-EpCAM staining. In experiments, suspended single cells were aspirated through microfluidic constriction channels with raw impedance data quantified and translated to Cspecific membrane and σcytoplasm. As to experimental results, significant differences in Cspecific membrane were observed for both live and fixed PC-3 cells with and without membrane staining, indicating that membrane staining proteins can contribute to electrical properties of cellular membranes. In addition, a significant decrease in σcytoplasm was located for PC-3 cells with and without fixation, suggesting that cytoplasm protein crosslinking during the fixation process can alter the cytoplasm conductivity. Overall, we have demonstrated how to classify single cells based on cellular electrical properties.

  13. Particle-rich cytoplasmic structure (PaCS): identification, natural history, role in cell biology and pathology.

    Science.gov (United States)

    Solcia, Enrico; Sommi, Patrizia; Necchi, Vittorio; Vitali, Agostina; Manca, Rachele; Ricci, Vittorio

    2014-09-22

    Cytoplasmic structures showing a selective concentration of both polyubiquitinated proteins and proteasome have been described in various epithelial, hematopoietic, mesenchymal and neural cells in vitro or in fetal tissues, as well as in chronically-infected, mutated preneoplastic and neoplastic tissues. These cytoplasmic structures differ from other ubiquitin-reactive cytoplasmic bodies, like sequestosomes, aggresome-like-induced structures in dendritic cells (DALIS)/non-dendritic cells (ALIS) and aggresomes in showing distinctive ultrastructural organization (particle-rich cytoplasmic structure or PaCS), a cytochemical pattern and a functional profile. Their formation can be induced in vitro in dendritic or natural killer cells by trophic factors and interleukin treatment. They originate in close connection with ribosomes, while, as a result of their growth, the cytoskeleton and other surrounding organelles are usually dislocated outside their core. Interestingly, these particulate cytoplasmic structures are often found to fill cytoplasmic blebs forming proteasome- and polyubiquitinated protein-discharging vesicles, called ectosomes, which are found to detach from the cell and freely float in the extracellular space. To clearly point out the importance of the polyubiquitinated proteins and proteasome containing cytoplasmic structures, their role in cell biology and pathology has been carefully analyzed.

  14. Production and Screening of Monoclonal Peptide Antibodies.

    Science.gov (United States)

    Trier, Nicole Hartwig; Mortensen, Anne; Schiolborg, Annette; Friis, Tina

    2015-01-01

    Hybridoma technology is a remarkable and indispensable tool for generating high-quality monoclonal antibodies. Hybridoma-derived monoclonal antibodies not only serve as powerful research and diagnostic reagents, but have also emerged as the most rapidly expanding class of therapeutic biologicals. In this chapter, an overview of hybridoma technology and the laboratory procedures used routinely for hybridoma production and antibody screening are presented, including characterization of peptide antibodies.

  15. Virus Strain Discrimination Using Recombinant Antibodies

    OpenAIRE

    Boonham, N.; Barker, I.

    2002-01-01

    Most routine testing for plant viruses is currently carried out using monoclonal and polyclonal antibodies. Traditional methods of antibody production however can be time consuming and require the use of expensive cell culture facilities. Recombinant antibody technology however is starting to make an impact in this area, enabling the selection of antibody fragments in a few weeks compared with the many months associated with traditional methods and requires only basic microbiological faciliti...

  16. Sequence and structural analysis of antibodies

    OpenAIRE

    Raghavan, A. K.

    2009-01-01

    The work presented in this thesis focusses on the sequence and structural analysis of antibodies and has fallen into three main areas. First I developed a method to assess how typical an antibody sequence is of the expressed human antibody repertoire. My hypothesis was that the more \\humanlike" an antibody sequence is (in other words how typical it is of the expressed human repertoire), the less likely it is to elicit an immune response when used in vivo in humans. In practi...

  17. A novel nucleo-cytoplasmic hybrid clone formed via androgenesis in polyploid gibel carp

    Directory of Open Access Journals (Sweden)

    Zhou Li

    2011-03-01

    Full Text Available Abstract Background Unisexual vertebrates have been demonstrated to reproduce by gynogenesis, hybridogenesis, parthenogenesis, or kleptogenesis, however, it is uncertain how the reproduction mode contributes to the clonal diversity. Recently, polyploid gibel carp has been revealed to possess coexisting dual modes of unisexual gynogenesis and sexual reproduction and to have numerous various clones. Using sexual reproduction mating between clone D female and clone A male and subsequent 7 generation multiplying of unisexual gynogenesis, we have created a novel clone strain with more than several hundred millions of individuals. Here, we attempt to identify genetic background of the novel clone and to explore the significant implication for clonal diversity contribution. Methods Several nuclear genome markers and one cytoplasmic marker, the mitochondrial genome sequence, were used to identify the genetic organization of the randomly sampled individuals from different generations of the novel clone. Results Chromosome number, Cot-1 repetitive DNA banded karyotype, microsatellite patterns, AFLP profiles and transferrin alleles uniformly indicated that nuclear genome of the novel clone is identical to that of clone A, and significantly different from that of clone D. However, the cytoplasmic marker, its complete mtDNA genome sequence, is same to that of clone D, and different from that of clone A. Conclusions The present data indicate that the novel clone is a nucleo-cytoplasmic hybrid between the known clones A and D, because it originates from the offspring of gonochoristic sexual reproduction mating between clone D female and clone A male, and contains an entire nuclear genome from the paternal clone A and a mtDNA genome (cytoplasm from the maternal clone D. It is suggested to arise via androgenesis by a mechanism of ploidy doubling of clone A sperm in clone D ooplasm through inhibiting the first mitotic division. Significantly, the selected nucleo-cytoplasmic

  18. Coordinated movement of cytoplasmic and transmembrane domains of RyR1 upon gating.

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    Montserrat Samsó

    2009-04-01

    Full Text Available Ryanodine receptor type 1 (RyR1 produces spatially and temporally defined Ca2+ signals in several cell types. How signals received in the cytoplasmic domain are transmitted to the ion gate and how the channel gates are unknown. We used EGTA or neuroactive PCB 95 to stabilize the full closed or open states of RyR1. Single-channel measurements in the presence of FKBP12 indicate that PCB 95 inverts the thermodynamic stability of RyR1 and locks it in a long-lived open state whose unitary current is indistinguishable from the native open state. We analyzed two datasets of 15,625 and 18,527 frozen-hydrated RyR1-FKBP12 particles in the closed and open conformations, respectively, by cryo-electron microscopy. Their corresponding three-dimensional structures at 10.2 A resolution refine the structure surrounding the ion pathway previously identified in the closed conformation: two right-handed bundles emerging from the putative ion gate (the cytoplasmic "inner branches" and the transmembrane "inner helices". Furthermore, six of the identifiable transmembrane segments of RyR1 have similar organization to those of the mammalian Kv1.2 potassium channel. Upon gating, the distal cytoplasmic domains move towards the transmembrane domain while the central cytoplasmic domains move away from it, and also away from the 4-fold axis. Along the ion pathway, precise relocation of the inner helices and inner branches results in an approximately 4 A diameter increase of the ion gate. Whereas the inner helices of the K+ channels and of the RyR1 channel cross-correlate best with their corresponding open/closed states, the cytoplasmic inner branches, which are not observed in the K+ channels, appear to have at least as important a role as the inner helices for RyR1 gating. We propose a theoretical model whereby the inner helices, the inner branches, and the h1 densities together create an efficient novel gating mechanism for channel opening by relaxing two right

  19. Immunoglobulin G4: an odd antibody

    NARCIS (Netherlands)

    R.C. Aalberse; S.O. Stapel; J. Schuurman; T. Rispens

    2009-01-01

    Despite its well-known association with IgE-mediated allergy, IgG4 antibodies still have several poorly understood characteristics. IgG4 is a very dynamic antibody: the antibody is involved in a continuous process of half-molecules (i.e. a heavy and attached light-chain) exchange. This process, also

  20. Anti-DNA antibodies in SLE

    Energy Technology Data Exchange (ETDEWEB)

    Voss, E.W.

    1988-01-01

    This book contains 8 chapters. Some of the titles are: Anti-DNA Antibodies in SLE: Historical Perspective; Specificity of Anti-DNA Antibodies in Systemic Lupus Erythematosus; Monoclonial Autoimmune Anti-DNA Antibodies; and Structure--Function Analyses of Anti-DNA Autoantibodies.

  1. Conditional expression of full-length humanized anti-prion protein antibodies in Chinese hamster ovary cells.

    Science.gov (United States)

    Mueller, Daniel A; Heinig, Lars; Ramljak, Sanja; Krueger, Astrid; Schulte, Reiner; Wrede, Arne; Stuke, Andreas W

    2010-12-01

    Because of their high antigen specificity and metabolic stability, genetically engineered human monoclonal antibodies are on the way to becoming one of the most promising medical diagnostics and therapeutics. In order to establish an in vitro system capable of producing such biosimilar antibodies, we used human constant chain sequences to design the novel human antibody expressing vector cassette pMAB-ABX. A bidirectional tetracycline (tet)-controllable promotor was used for harmonized expression of immunoglobulin type G (IgG) heavy and light chains. As an example we used anti-prion protein (anti-PrP) IgGs. Therefore, the variable heavy (V(H)) and light chain (V(L)) sequences of anti-PrP antibodies, previously generated in our laboratory by DNA immunization of prion protein knock-out mice, were isolated from murine hybridoma cell lines and inserted into pMAB-ABX vector. After transfection of Chinese hamster ovary (CHO) cells, a number of stable antibody producing cell clones were selected. One cell line (pMAB-ABX-13F10/3B5) stably expressing the recombinant humanized antibody (rechuAb) 13F10/3B5 was selected for detailed characterization by Western blot, immunofluorescence, and flow cytometric analyses. The full-length recombinant humanized IgG antibody showed a high level of expression in the cytoplasm. In conclusion, the new cell system described here is a suitable tool to produce functional intact full-length humanized IgG antibodies. PMID:21087094

  2. Isolation of human monoclonal antibodies to the envelope e2 protein of hepatitis C virus and their characterization.

    Directory of Open Access Journals (Sweden)

    Yohko K Shimizu

    Full Text Available We isolated and characterized two human monoclonal antibodies to the envelope E2 protein of hepatitis C virus (HCV. Lymphoblastoid cell lines stably producing antibodies were obtained by immortalizing peripheral blood mononuclear cells of a patient with chronic hepatitis C using Epstein-Barr virus. Screening for antibody-positive clones was carried out by immunofluorescence with Huh7 cells expressing the E2 protein of HCV strain H (genotype 1a isolated from the same patient. Isotype of resulting antibodies, #37 and #55, was IgG1/kappa and IgG1/lambda, respectively. Epitope mapping revealed that #37 and #55 recognize conformational epitopes spanning amino acids 429 to 652 and 508 to 607, respectively. By immunofluorescence using virus-infected Huh7.5 cells as targets both antibodies were reactive with all of the nine different HCV genotypes/subtypes tested. The antibodies showed a different pattern of immuno-staining; while #37 gave granular reactions mostly located in the periphery of the nucleus, #55 gave diffuse staining throughout the cytoplasm. Both antibodies were shown by immuno-gold electron microscopy to bind to intact viral particles. In a neutralization assay (focus-forming unit reduction using chimeric infectious HCV containing structural proteins derived from genotypes 1a, 1b, 2a, 2b, 3a, 4a, 5a, 6a, and 7a, #55 inhibited the infection of all HCV genotypes tested but genotype 7a to a lesser extent. #37 did not neutralize any of these viruses. As a broadly cross-neutralizing human antibody, #55 may be useful for passive immunotherapy of HCV infection.

  3. Susceptibility to virus-cell fusion at the plasma membrane is reduced through expression of HIV gp41 cytoplasmic domains.

    Science.gov (United States)

    Malinowsky, Katharina; Luksza, Julia; Dittmar, Matthias T

    2008-06-20

    The cytoplasmic tail of the HIV transmembrane protein plays an important role in viral infection. In this study we analyzed the role of retroviral cytoplasmic tails in modulating the cytoskeleton and interfering with virus-cell fusion. HeLaP4 cells expressing different HIV cytoplasmic tail constructs showed reduced acetylated tubulin levels whereas the cytoplasmic tail of MLV did not alter microtubule stability indicating a unique function for the lentiviral cytoplasmic tail. The effect on tubulin is mediated through the membrane proximal region of the HIV cytoplasmic tail and was independent of membrane localization. Site-directed mutagenesis identified three motifs in the HIV-2 cytoplasmic tail required to effect the reduction in acetylated tubulin. Both the YxxPhi domain and amino acids 21 to 45 of the HIV-2 cytoplasmic tail need to be present to change the level of acetylated tubulin in transfected cells. T-cells stably expressing one HIV-2 cytoplasmic tail derived construct showed also a reduction in acetylated tubulin thus confirming the importance of this effect not only for HeLaP4 and 293T cells. Challenge experiments using transiently transfected HeLaP4 cells and T cells stably expressing an HIV cytoplasmic tail construct revealed both reduced virus-cell fusion and replication of HIV-1(NL4.3) compared to control cells. In the virus-cell fusion assay only virions pseudotyped with either HIV or MLV envelopes showed reduced fusion efficiency, whereas VSV-G pseudotyped virions where not affected by the expression of HIV derived cytoplasmic tail constructs, indicating that fusion at the plasma but not endosomal membrane is affected. Overexpression of human histone-deacetylase 6 (HDAC6) and constitutively active RhoA resulted in a reduction of acetylated tubulin and reduced virus-cell fusion as significant as that observed following expression of HIV cytoplasmic tail constructs. Inhibition of HDAC6 showed a strong increase in acetylated tubulin and increase of

  4. Eosinophilic granulomatosis with polyangiitis (Churg-Strauss syndrome and pulmonary thromboembolism: an overlooked concomitance

    Directory of Open Access Journals (Sweden)

    Vilma Takayasu

    2013-06-01

    Full Text Available The Eosinophilic Granulomatosis with Polyangiitis (formerly Churg- Strauss Syndrome (EGPA is a systemic inflammatory disease characterized by the presence of rhinitis, asthma, peripheral eosinophilia, and vasculitis—the latter being characteristic of the late stage of the disease. After several years from the onset of the disease, small- and medium-sized vessel vasculitis ensues, undertaking various organs and systems. Upper and lower airways, skin, nervous system, gastrointestinal tract, heart, and kidneys are the most commonly involved organs. It is believed that tissue injury is the result of processes mediated by antineutrophil cytoplasmic antibody (ANCA, or toxic mediators released by eosinophils. Although it is classified as ANCA-associated vasculitis, these autoantibodies are present in only 40% of cases. The authors report the case of a patient with EGPA, who had a history of asthma, peripheral and central neuropathy, palpable purpura, gastrointestinal micro perforation, peripheral eosinophilia, and the presence of myeloperoxidase-antineutrophil cytoplasmic antibody. Inflammatory parameters improved after the initiation of treatment, but 1 month after hospital discharge the patient developed symptoms compatible with pulmonary embolism and died. Thrombophilia that occurs in EGPA is due to the interaction between the inflammatory response and eosinophilia with the clotting system resulting in a pro-thrombotic state. Although not yet well-determined, the authors call attention to the possibility of the impact of thromboembolic events on the prognosis of patients with EGPA. In addition to the adequate immunosuppressive treatment, prophylaxis and treatment for thrombosis should never be overlooked.

  5. Pulmonary hyalinizing granuloma. A limited form of Wegener's granulomatosis?

    Science.gov (United States)

    Gorini, M; Forloni, F; Pezzoli, A; Pezzica, E

    1998-01-01

    Pulmonary hyalinizing granuloma is an uncommon disease that consists of slowly enlarging nodules in the pulmonary parenchyma. It occurs rarely: in fact, fewer than 70 case reports have been published in the past 20 years. It is important however in the differential diagnosis of lung diseases manifesting multiple pulmonary nodules. The etiology and pathogenesis of this disorder are unknown. Evidence suggests that the nodules could be the result of a chronic exaggerated immune response to infectious agents or to any other process in which antigen-antibody complexes are involved. More than 50% of the patients reported have evidence of autoimmune phenomena, e.g. positive antinuclear antibodies, a positive rheumatoid factor, or circulating immune complexes. The present report describes, for the first time, a case of pulmonary hyalinizing granuloma in which the patient had antineutrophil cytoplasmic autoantibodies with a granular cytoplasmatic pattern with typical central accentuation of fluorescence intensity and negative nuclei. The presence of antineutrophil cytoplasmic autoantibodies suggests that pulmonary hyalinizing granuloma could be regarded as a localized, non-evolving, form of Wegener's granulomatosis or a purely granulomatous Wegener's granulomatosis. PMID:9859576

  6. Dengue fever triggering systemic lupus erythematosus and lupus nephritis: a case report

    Directory of Open Access Journals (Sweden)

    Talib SH

    2013-10-01

    Full Text Available SH Talib, SR Bhattu, R Bhattu, SG Deshpande, DB Dahiphale Department of Medicine and Nephrology, MGM Medical College and Hospital, Aurangabad, Maharashtra, India Abstract: We report a rare case of dengue fever triggering systemic lupus erythematosus and lupus nephritis. The patient presented herself during a large outbreak of dengue fever in December 2012 in Maharashtra, India. The diagnosis of dengue fever was confirmed by the presence of NS-1 antigen during the first few days of febrile illness. Eight weeks later, kidney tissue biopsy studies revealed evidence of lupus nephritis on microscopic examination and immunofluorescence. The report interpreted it as focal proliferative glomerulonephritis and segmental sclerosis (Stage IIIC. The case was also found positive for perinuclear antineutrophil cytoplasmic antibodies by indirect immunofluorescence assay. An active and effective management of a case essentially calls for clear perception of differentiating dengue-induced lupus flare, antineutrophil cytoplasmic antibody-related nephropathy, and/or dengue-induced de-novo lupus disease. Dengue viremia may be the trigger for immune complex formation in patients who are predisposed to developing autoimmune diseases. The present case explains the importance of considering the diagnosis of dengue-related lupus nephritis as an atypical occurrence in appropriate situations, as in this case. It would not be improper to regard this escalating disease as an expanded feature of dengue. Keywords: kidney biopsy, glomerulonephritis, segmental sclerosis, lupus flare, dengue viremia, autoimmune, de-novo lupus nephritis

  7. Granulomatosis with polyangiitis: rapidly progressive necrotizing glomerulonephritis in a pediatric patient

    Directory of Open Access Journals (Sweden)

    Luna M

    2014-04-01

    Full Text Available Mariana Luna,1 Victoria Bocanegra,3 Patricia G Vallés1,21Nephrology Division, Pediatric Department, Dr Humberto Notti Pediatric Hospital, Mendoza, Argentina; 2Pathophysiology Area, Pathology Department, School of Medicine, National Cuyo University, Mendoza, Argentina; 3National Council of Scientific and Technical Research of Argentina (CONICET, Buenos Aires, ArgentinaAbstract: Granulomatosis with polyangiitis (GPA is associated with a broad range of clinical manifestations including renal disease. It is a systemic vasculitis that is rarely encountered in children. We present a 14-year-old girl who suffered from pharyngitis 1 week before admittance to hospital. She was admitted for macroscopic hematuria and oliguria, under the possibility of nephritic syndrome. Renal failure with rapidly progressive glomerulonephritis occurred within 24 hours. Immunologic tests showed the presence of type-C anti-neutrophil cytoplasmic antibodies (c-ANCA with antiproteinase 3 specificity and renal biopsy revealed pauci-immune crescentic focal necrotizing glomerulonephritis. Treatment including methylprednisolone and cyclophosphamide intravenous pulses allowed renal recovery after 3 weeks. The clinical, hematological, and biochemical parameters improved substantially, achieving remission. Granulomatosis with polyangiitis, although rare in children, should be considered in the above clinical scenario. This case underlines that knowledge of renal histology diagnosis and early aggressive immunosuppressive therapy are essential for the management of these patients.Keywords: acute renal failure, vasculitis, crescentic pauci-immune glomerulonephritis-Type-C, antineutrophil cytoplasmic antibodies (c-ANCA-macroscopic hematuria

  8. The role of antibodies in myasthenia gravis.

    Science.gov (United States)

    De Baets, M; Stassen, M H W

    2002-10-15

    Myasthenia gravis is an autoimmune disease associated with antibodies directed to the postsynaptic acetylcholine receptor. These antibodies reduce the number of receptors. Autoantibodies against AChR and other muscle antigens can be used for the diagnosis of myasthenia gravis and related disorders. The origin and the role of these antibodies in the disease are discussed. Experimental autoimmune myasthenia gravis, an experimental model closely mimicking the disease, has provided answers to many questions about the role of antibodies, complement macrophages and AChR anchor proteins. Genetically modified anti-AChR antibodies may also be used in the future to treat myasthenia. PMID:12220686

  9. Production of Monoclonal Antibody against Human Nestin.

    Science.gov (United States)

    Hadavi, Reza; Zarnani, Amir Hassan; Ahmadvand, Negah; Mahmoudi, Ahmad Reza; Bayat, Ali Ahmad; Mahmoudian, Jafar; Sadeghi, Mohammad-Reza; Soltanghoraee, Haleh; Akhondi, Mohammad Mehdi; Tarahomi, Majid; Jeddi-Tehrani, Mahmood; Rabbani, Hodjattallah

    2010-04-01

    We have employed a peptide-based antibody generation protocol for producing antibody against human nestin. Using a 12-mer synthetic peptide from repetitive region of human nestin protein devoid of any N- or O-glyco-sylation sequences, we generated a mouse monoclonal antibody capable of recognizing human, mouse, bovine, and rat nestin. A wide variety of nestin proteins ranging from 140-250 kDa was detected by this antibody. This antibody is highly specific and functional in applications such as ELISA, flow cytometry, immunocytochemistry, and Western blot assays. PMID:23407796

  10. Production of Monoclonal Antibody against Human Nestin

    OpenAIRE

    Hadavi, Reza; Zarnani, Amir Hassan; Ahmadvand, Negah; Mahmoudi, Ahmad Reza; Bayat, Ali Ahmad; Mahmoudian, Jafar; Sadeghi, Mohammad-Reza; Soltanghoraee, Haleh; Akhondi, Mohammad mehdi; Tarahomi, Majid; Jeddi-Tehrani, Mahmood; Rabbani, Hodjattallah

    2010-01-01

    We have employed a peptide-based antibody generation protocol for producing antibody against human nestin. Using a 12-mer synthetic peptide from repetitive region of human nestin protein devoid of any N- or O-glyco-sylation sequences, we generated a mouse monoclonal antibody capable of recognizing human, mouse, bovine, and rat nestin. A wide variety of nestin proteins ranging from 140–250 kDa was detected by this antibody. This antibody is highly specific and functional in applications such a...

  11. Monoclonal Antibody Therapies against Anthrax

    OpenAIRE

    Zhaochun Chen; Mahtab Moayeri; Robert Purcell

    2011-01-01

    Anthrax is a highly lethal infectious disease caused by the spore-forming bacterium Bacillus anthracis. It not only causes natural infection in humans but also poses a great threat as an emerging bioterror agent. The lethality of anthrax is primarily attributed to the two major virulence factors: toxins and capsule. An extensive effort has been made to generate therapeutically useful monoclonal antibodies to each of the virulence components: protective antigen (PA), lethal factor (LF) and ede...

  12. Antibody Peptide Based Antifungal Immunotherapy

    OpenAIRE

    Magliani, Walter; Conti, Stefania; Giovati, Laura; Zanello, Pier Paolo; Sperindè, Martina; Ciociola, Tecla; Polonelli, Luciano

    2012-01-01

    Fungal infections still represent relevant human illnesses worldwide and some are accompanied by unacceptably high mortality rates. The limited current availability of effective and safe antifungal agents makes the development of new drugs and approaches of antifungal vaccination/immunotherapy every day more needed. Among them, small antibody(Ab)-derived peptides are arousing great expectations as new potential antifungal agents. In this topic, the search path from the study of the yeast kill...

  13. Dehydration-induced redistribution of amphiphilic molecules between cytoplasm and lipids is associated with desiccation tolerance in seeds

    NARCIS (Netherlands)

    Buitink, J.; Leprince, O.; Hoekstra, F.A.

    2000-01-01

    This study establishes a relationship between desiccation tolerance and the transfer of amphiphilic molecules from the cytoplasm into lipids during drying, using electron paramagnetic resonance spectroscopy of amphiphilic spin probes introduced into imbibed radicles of pea (Pisum sativum) and cucumb

  14. Screening and Identifying Two Specific Molecular Markers in Maintainer Line of Cytoplasmic Male Sterility Cauliflower (Brassica oleracea var.botrytis)

    Institute of Scientific and Technical Information of China (English)

    C.G. Wang; J.M. Gao; H. Li; Y. Gu; X.Q. Chen; W.Q. Song

    2007-01-01

    @@ Cytoplasmic male sterility (CMS) is a maternally inherited trait that prevents the production of function pollen, but maintains female fertility. It has been widely used in breeding programs to product F1 hybrid seed in some crops (Hanson, 1991).

  15. Regulation of adherence and virulence by the Entamoeba histolytica lectin cytoplasmic domain, which contains a beta2 integrin motif.

    Science.gov (United States)

    Vines, R R; Ramakrishnan, G; Rogers, J B; Lockhart, L A; Mann, B J; Petri, W A

    1998-08-01

    Killing of human cells by the parasite Entamoeba histolytica requires adherence via an amebic cell surface lectin. Lectin activity in the parasite is regulated by inside-out signaling. The lectin cytoplasmic domain has sequence identity with a region of the beta2 integrin cytoplasmic tail implicated in regulation of integrin-mediated adhesion. Intracellular expression of a fusion protein containing the cytoplasmic domain of the lectin has a dominant negative effect on extracellular lectin-mediated cell adherence. Mutation of the integrin-like sequence abrogates the dominant negative effect. Amebae expressing the dominant negative mutant are less virulent in an animal model of amebiasis. These results suggest that inside-out signaling via the lectin cytoplasmic domain may control the extracellular adhesive activity of the amebic lectin and provide in vivo demonstration of the lectin's role in virulence.

  16. Application of Monoclonal Antibodies in Veterinary Parasitology

    Directory of Open Access Journals (Sweden)

    Gupta A.

    2011-08-01

    Full Text Available The discovery of hybridoma technology by Kohler and Milstein in 1975, heralded a new era in antibody research. Mouse hybridomas were the first reliable source of monoclonal antibodies. The generation of monoclonal antibodies from species other than rats and mice, has developed slowly over the last 30 years. The advent of antibody engineering and realization of the advantages of non murine antibodies has increased their relevance recently. However, in the area of veterinary parasitology, monoclonal antibodies are just beginning to fulfill the promises inherent in their great specificity for recognizing and selectively binding to antigens. This review describes the recent advances in the application of monoclonal antibodies for immunodiagnosis / prophylaxis and immunotherapy of parasitic diseases. [Vet. World 2011; 4(4.000: 183-188

  17. Compartmentalization of membrane trafficking, glucose transport, glycolysis, actin, tubulin and the proteasome in the cytoplasmic droplet/Hermes body of epididymal sperm.

    Science.gov (United States)

    Au, Catherine E; Hermo, Louis; Byrne, Elliot; Smirle, Jeffrey; Fazel, Ali; Kearney, Robert E; Smith, Charles E; Vali, Hojatollah; Fernandez-Rodriguez, Julia; Simon, Paul H G; Mandato, Craig; Nilsson, Tommy; Bergeron, John J M

    2015-08-01

    Discovered in 1909 by Retzius and described mainly by morphology, the cytoplasmic droplet of sperm (renamed here the Hermes body) is conserved among all mammalian species but largely undefined at the molecular level. Tandem mass spectrometry of the isolated Hermes body from rat epididymal sperm characterized 1511 proteins, 43 of which were localized to the structure in situ by light microscopy and two by quantitative electron microscopy localization. Glucose transporter 3 (GLUT-3) glycolytic enzymes, selected membrane traffic and cytoskeletal proteins were highly abundant and concentrated in the Hermes body. By electron microscope gold antibody labelling, the Golgi trafficking protein TMED7/p27 localized to unstacked flattened cisternae of the Hermes body, as did GLUT-3, the most abundant protein. Its biogenesis was deduced through the mapping of protein expression for all 43 proteins during male germ cell differentiation in the testis. It is at the terminal step 19 of spermiogenesis that the 43 characteristic proteins accumulated in the nascent Hermes body. PMID:26311421

  18. Ro small cytoplasmic ribonucleoproteins are a subclass of La ribonucleoproteins: Further characterization of the Ro and La small ribonucleoproteins from uninfected mammalian cells

    Energy Technology Data Exchange (ETDEWEB)

    Hendrick, J.P.; Wolin, S.L.; Rinke, J.; Lerner, M.R.; Steitz, J.A.

    1981-12-01

    Small ribonucleic acid (RNA)-protein complexes precipitated by anti-Ro and anti-La antibodies from lupus patients have been examined with emphasis on their RNA components. In both ribonucleoprotein (RNP) classes, the numbers of different RNA molecules and their sequences vary between mouse and human cells. The complex mixtures of La RNAs include two previously sequenced 4.5S RNAs from mouse cells and 5S ribosomal RNA-like molecules from both mouse and human cells. All Ro and La RNAs possess 5'-triphosphates. Some La RNAs have internal modifications typical of transfer RNAs. The RoRNPs are quite stable and are localized by immunofluorescence in the cell cytoplasm, whereas the majority of the La RNPs turn over rapidly and reside in the nucleus. Despite these differences, reconstitution experiments show that the Ro particles carry the La as well as the Ro determinant. Studies using a nuclear transcription system demonstrate that most of the La RNAs are synthesized by RNA polymerase III. The possibility that the La protein(s) functions in the transcription or maturation of all RNA polymerase III transcripts is discussed.

  19. Classification of Cells with Membrane Staining and/or Fixation Based on Cellular Specific Membrane Capacitance and Cytoplasm Conductivity

    OpenAIRE

    Song-Bin Huang; Yang Zhao; Deyong Chen; Shing-Lun Liu; Yana Luo; Tzu-Keng Chiu; Junbo Wang; Jian Chen; Min-Hsien Wu

    2015-01-01

    Single-cell electrical properties (e.g., specific membrane capacitance (Cspecific membrane) and cytoplasm conductivity (σcytoplasm)) have been regarded as potential label-free biophysical markers for the evaluation of cellular status. However, whether there exist correlations between these biophysical markers and cellular status (e.g., membrane-associate protein expression) is still unknown. To further validate the utility of single-cell electrical properties in cell type classification, Cspe...

  20. Effects of Nuclear Genomes on Anther Development in Cytoplasmic Male Sterile Chicories (Cichorium intybus L.): Morphological Analysis

    OpenAIRE

    Ildephonse Habarugira; Theo Hendriks; Marie-Christine Quillet; Jean-Louis Hilbert; Caroline Rambaud

    2015-01-01

    The Cichorium intybus flower development in fertile, cytoplasmic male sterility (CMS 524) and various phenotypes carrying the 524 male sterile cytoplasm was investigated macroscopically and by light microscopy. The development was similar in fertile and in male sterile florets up to meiosis, and then it was affected in anther wall structure and pollen grain development in male sterile floret. In the male sterile plants, the tapetum intrusion after meiosis was less remarkable, the microspores...