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Abstract.Anti-Sm antibodies recognize a group of small, nuclear RNA-protein complexes (snRNPs) containing U 1, U2, U4, U5, and U6 snRNAs. Anti-RNP antibodies only react with U I snRNA-containing complexes.The intranuclear distribution of snRNP particles was studied by double immunofluorescence staining of human fibroblasts. Mouse monoclonal anti-Sm antibodies and polyclonal patient sera reacting with different peptides in the snRNP complexes were used. The immunofluorescence patterns obtained with fluorescein isothiocyanate-conjugated anti-mouse Ig and tetramethylrhodamine isothiocyanate-conjugated anti-human Ig second antibodies were examined using computer analysis of digitized images. With this approach the similarity of different patterns could be visualized and estimated with mathematical methods. It was found that human anti-Sm serum as well as three different anti-RNP sera produced speckled patterns overlapping with the anti-Sm monoclonal pattern. Thus, Sm antigenic intranuclear domains also reacted with anti-RNP antibodies, suggesting a high degree of co-localization of the antigenic structures. A partial overlap was found between speckles detected by mouse anti-Sm antibodies and a human La-antiserum. No significant co-localization occurred between speckles detected by mouse anti-Sm antibodies and speckles detected by human antisera reacting with Scl-70 and centromeric antigens.As the U 1 snRNP complex is believed to play a role in the splicing of RNA polymerase II transcripts, it appears that the speckles detected by Sm and RNP antibodies may be regions of hnRNA synthesis and m.RNA processing. Although no function has been demonstrated for the U2, U4, U5, and U6 snRNPs, the co-localization with the U 1 RNA complexes shown in this report indicate that they too participate in some aspect of mRNA processing. The results suggest that computer-assisted analysis of nuclear immunofluorescence patterns will be a useful tool in studies of the spatial and functional organization of the interphase nucleus. IMMUNE staining of mammalian cells with autoimmune sera frequently results in a speckled nuclear pattern. The two major specificities that produce speckled patterns are known as Sm and RNP. These autoantibodies occur primarily in systemic lupus erythematosus, mixed connective tissue disease, and less frequently in other autoimmune diseases (3,31,35).The Sm and RNP antigenic structures are thus concentrated in certain nuclear domains. Immunochemical and biochemical studies have elucidated the nature of the antigens (1, 11-13, 16, 18, 20, 25, 33, 34, 37, 38). Sm-antibodies were shown to immunoprecipitate RNA-protein complexes containing a class of small nuclear RNAs designated U l, U2, U4, U5, and U6 snRNAs) RNP-antibodies, on the other hand, immunoprecipitated only U 1 RNA-containing complexes (18). Proteins were required for antigenicity (18). The fact that small, U3 is nucleolar and not precipitated by Sm/RNP antibodies. nuclear RNA-protein complex (snRNP) 2 particles containing only U l RNA can be immunoprecip...

Recent experimental data demonstrate cardiovascular effects of the GH secretagogues (GHSs) hexarelin and ghrelin, the proposed natural ligand for the GHS receptor. Moreover, specific cardiac binding sites for GHSs have been suggested. The aim of the present study was to investigate if the natural ligand ghrelin and synthetic GHS peptide hexarelin and analogues have direct effects on the cardiomyocyte cell line, H9c2. Hexarelin stimulated thymidine incorporation in a dose-dependent manner with significant responses at 3 µM (147 3% of control, P<0·01) and elicited maximal effects at concentrations around 30 µM. This activity was seen already after 12 h of incubation with a maximal effect after 18 h (176 9% of control, P<0·01). Ghrelin also had a significant stimulatory effect on thymidine incorporation (129 2% of control at 3 µM and 18 h, P<0·05). The stimulatory effect on thymidine incorporation of hexarelin, Tyr-Ala-hexarelin, EP80317 and ghrelin was specific and no stimulatory effect was observed with the truncated GH-releasing peptide EP51389 or the non-peptidyl GHS MK-0677. In competitive binding studies, 125 I-labeled Tyr-Alahexarelin was used as radioligand and competition curves showed displacement with hexarelin, Tyr-Ala-hexarelin, EP80317 and ghrelin, whereas MK-0677 and EP51389 produced very little displacement at 1 µM concentration, adding further support for an alternative subtype binding site in the heart compared with the pituitary. In conclusion, we have demonstrated a dose-dependent and specific stimulation of cardiomyocyte thymidine incorporation by natural and synthetic GHS analogues, suggesting increased cell proliferation and binding of GHS to H9c2 cardiomyocyte cell membranes. These findings support potential peripheral effects of GHS on the cardiovascular system independent of an increased GH secretion.

Antibodies to ribonucleoproteins (RNP) and to the Sni antigen in sera from patients with mixed connective tissue disease (MCTD) and systemic lupus erythematosus were studied using the techniques of immunioblotting and immunoprecipitation of U small nuclear RNPs. A cross-sectional study indicated that antibodies reacting with a 68K protein were associated with anti-RNP specificity in MCTD, but rarely occurred in systemic lupus erythematosus patients' sera. A longitudinal study demonstrated the persistence of MCTD blotting patterns over many years, and the subsequent disappearance of those specificities in sera from patients who were in prolonged remission.Patients with connective tissue diseases produce antibodies which react with various cellular components such as DNA, RNA, proteins, and _____ From the Karolinska Institute, Stockholm, Sweden and the University of MissouriXolumbia.