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Biomedical subjects

S A Rich

Publications and source records attributed to S A Rich.

At least 19 recordsLinked to original sources
PubMed1996

Purification, microsequencing, and immunolocalization of p36, a new interferon-alpha-induced protein that is associated with human lupus inclusions.

The trace interferon-alpha-induced protein, p36, was induced in Raji cells in association with lupus inclusions. It was solubilized in a nonionic detergent buffer, enriched by differential centrifugation and by preparative isoelectric focusing, and purified to homogeneity on two-dimensional protein gels. Failure to obtain N-terminal amino acid sequence, however, suggested a blocked alpha-amino group. Sequences of six tryptic peptides, 13-19 amino acids in length, were obtained after digestion, microbore-high performance liquid chromotography purification, and chemical sequence analysis. None of the six sequences, which represented approximately 25% of the entire protein, shared any meaningful homologies with entries in protein sequence repositories. Raji-cell p36 was shown in Western blots with antipeptide antibodies to be induced at least 400-fold and by immunofluorescence microscopy to co-localize with the endoplasmic reticulum resident protein, protein disulfide isomerase. These results show that p36 is a new interferon-alpha-induced protein that localizes in the endoplasmic reticulum, the cell region in which the lupus inclusions form, and that p36 is probably physically associated with the lupus inclusions.

Amino Acid Sequence↗
PubMed1996

Interferon-alpha-induced human lupus inclusions and p36 protein in cancer and AIDS.

Previous work showed that IFN-alpha induced the autoimmune-associated lupus inclusions (LI) in all 16 umbilical cord mononuclear cell samples from healthy mothers. In contrast, IFN-alpha induced LI and the LI-associated protein, p36, in only 2 of 16 human B lymphoblastoid cell lines. Resistance of these 14 cell lines to form LI and p36 may be due to their stage of development or differentiation or their transformed state. We sought to determine whether aging, neoplastic transformation, and HIV infection affected the observed IFN-alpha induction of LI in cord blood mononuclear cells. Expression of LI and p36 was investigated in PBMC on IFN-alpha chemotherapy and on culturing IFN-alpha with PBMC samples prepared from healthy adults and AIDS patients. The IFN-alpha induction of LI (detected by electron microscopy) or p36 (detected by two-dimensional gels) in all of the PBMC samples from these individuals was indistinguishable from the cord blood mononuclear cell response. Furthermore, induction of p36 and LI was not a good indicator of effective IFN-alpha chemotherapy. It may be consequential for autoimmunity induced by IFN-alpha in cancer, AIDS, and systemic lupus erythematosus (SLE). An essential biologic role for p36 and LI is suggested by a highly homologous p36 gene in the invertebrate Caenorhabditis elegans.

Acquired Immunodeficiency Syndrome↗
PubMed1995

De novo synthesis and secretion of a 36-kD protein by cells that form lupus inclusions in response to alpha-interferon.

In response to the pure recombinant human alpha-IFN, IFLrA, Raji and Daudi were the only two cell lines among 19 human lymphoblastoid cell lines tested that formed the human lupus inclusions (LI) to a high frequency. Raji, Daudi, and five other cell lines were examined for protein changes that might accompany LI formation. Their selection was based upon T or B origin, association with Epstein-Barr virus, and ability to form LI. A trace protein of an estimated molecular mass of 36 kD (p36) and an isoelectric point of 5.6 was detected on two-dimensional gels only of alpha-IFN-treated Raji and Daudi cells. Gamma-IFN did not induce p36 or LI in any of these seven cell lines. In Daudi cells p36 and LI formed simultaneously in response to IFLrA, and persisted until the alpha-IFN-induced death of the culture. In Raji cells, p36 and LI appearance and disappearance coincided with the addition and removal of alpha-IFN. Fractionation of Raji cells with nonionic-detergent buffer placed p36 with the inclusions in the cytoplasmic supernatant. With detergent-free buffer p36 and LI were distributed evenly between the nuclear and cytoplasmic fractions. Pulse-chase experiments revealed that p36 was secreted. The de novo synthesis of p36 with alpha-IFN treatment was shown by labeling the cell proteins with [35S] methionine before and after the addition of alpha-IFN. These results along with previous results on the de novo synthesis of LI in the endoplasmic reticulum (which is involved in the processing and secretion of proteins) suggest a role for LI in the synthesis and secretion of p36.

B-Lymphocytes↗
PubMed1993

Treatment of foxglove extract poisoning with digoxin-specific Fab fragments.

A 22-year-old man presented to our emergency department after an intentional overdose of a homemade foxglove extract. Clinical symptoms with symptomatic bradyarrhythmia and ECG changes were consistent with cardiac glycoside poisoning. Treatment with digoxin-specific Fab fragments resulted in transient clinical and ECG improvement. Serum immunoassay demonstrated a digitoxin-like glycoside. The serum levels showed no evidence of altered elimination or distribution with Fab therapy despite temporary improvements in the clinical course. The use of Fab did not result in a shortened clinical course in this episode of foxglove poisoning, as one would expect in the setting of commercial glycoside product poisoning.

Adult↗
PubMed1990

Human lupus-type inclusions in umbilical cord blood mononuclear cells.

Mononuclear cells purified from umbilical cord blood activated oligoadenylate synthetase and formed human lupus-type inclusions (LI) when cultured with the purified recombinant human leukocyte interferon, IFLrA. LI frequencies increased from 0% to a low of 0.75% and a high of 6.25% in 4-day cultures with IFLrA (100 units/ml). These interferon-induced responses in the mononuclear cells of neonates indicate that LI are solely an intrinsic product of normal cells, and not an exogenous virus or some other environmental agent.

2',5'-Oligoadenylate Synthetase↗
PubMed1989

Evidence that the interferon-induced Daudi cell human lupus inclusions are de novo synthesized complexes of ribonucleoprotein and membrane.

Quantitative electron microscope autoradiography has been used to define the macromolecular composition of the interferon-induced human lupus-type inclusions (LI) in the human B lymphoblastoid cell line, Daudi. LI were first apparent in Daudi cell cultures 12 h after the addition of 100 units/ml of the purified recombinant human leukocyte interferon, IFLrA. Radiolabels were added at this time and allowed to incorporate over the following 12 h during which an estimated greater than 99% of the LI material present at 24 h was formed. The LI-incorporated radiolabels were present only during this discrete 12-h period after the interferon activation of LI cell pathways in order to detect LIs de novo synthesized macromolecular components. The estimate relative specific activities of the LI-incorporated radiolabels were: choline at 4.042, mannose at 2.631, uridine at 0.664, glucosamine at 0.578, and amino acids at 0.477. With thymidine the estimated LI specific activity was 0.000. LI isolated from whole cells retained the tubular elements and the interwoven membrane network. These results provide direct evidence that the interferon-induced Daudi cell LI are de novo synthesized complexes of ribonucleoprotein and membrane.

Amino Acids↗
PubMed1986

Induction of lupus inclusions by sera from patients with systemic lupus erythematosus.

A sensitive in vitro bioassay for the alpha-interferon induction of lupus-type inclusions (LI) has been established with the human B lymphoblastoid cell line, Daudi. Sera from 11 patients with systemic lupus erythematosus (SLE) were evaluated with this assay. LI induction by these sera increased in proportion to their antiviral activity on Madin-Darby bovine kidney (MDBK) cells. Two of these sera did not induce LI; they showed no antiviral activity on the MDBK cell assay. Clinically and serologically, their donors were in remission. Two sera induced the formation of LI that exceeded the maximum frequencies obtained with 3 alpha-interferon preparations. These sera had the greatest antiviral activities, and their donors had the greatest disease activities. Antisera to alpha-interferons prevented the induction of LI with the pure and homogeneous recombinant human leukocyte interferon, IFLrA, and SLE sera. Together, these results provide evidence that the alpha-interferon endogenous to SLE patients has a great ability to induce LI, and the SLE serum induction of LI corresponds well to the patient's disease activity.

Animals↗
PubMed1985

Remediation of memory disorders: experimental evaluation of the spaced-retrieval technique.

Research concerning remediation of memory disorders has frequently been concerned with mnemonic techniques that demand a great deal of elaborative and effortful processing. The present study examines a relatively simple technique, known as spaced retrieval, in which patients are taught to retrieve information at increasingly long temporal intervals after initial presentation. Results indicated that the spaced-retrieval technique aided patients' learning of new information. There was also evidence of learning to learn: Two of the four patients who were studied learned to use the technique in the absence of explicit cues from the experimenter. Issues pertaining to the possible usefulness of spaced retrieval in everyday life are discussed.

Adult↗
PubMed1984

Purified recombinant human leukocyte interferons IFLrA and IFLrD induce human lupus inclusions in Raji and Daudi cells.

Raji and Daudi are human B lymphoblastoid cell lines that readily form lupus inclusions (LIs; TRS) when grown in medium supplemented with leukocyte-, or fibroblast-derived interferon (IFN-alpha, -beta, respectively). WISH, MDBK, and GM2504 are three cell lines commonly used to measure antiviral activities. None of them form LIs in their antiviral response to alpha or immune (gamma)IFN. This distinguishes between the abilities of a cell to develop an antiviral state and to form LIs in response to IFN. Human (Hu) lymphoblastoid IFN and the two pure and homogeneous recombinant human IFN-alpha proteins IFLrA and IFLrD induce LIs in Raji cells and Daudi cells. In Daudi, a simultaneous inhibition of cell growth occurs. When compared by antiviral activities, IFLrA inhibits the growth of Daudi cells more, while IFLrD induces the greater frequency of LIs. According to molecular concentration, IFLrA and IFLrD at 133 X 10(-13) M induce LIs in Daudi cells to their maximum frequency. Growth inhibition for these same cell samples is also at maximum for IFLrA, but only 25% of maximum for IFLrD. Our results with Raji and Daudi cells provide evidence against a cause-and-effect relationship between these two biologic responses to IFN by Daudi cells. They also provide evidence for distinct, but interacting, intracellular pathways. This phenomenon is a new explanation for some of the biologic diversity shown for the HuIFNs-alpha.

B-Lymphocytes↗
PubMed1981

Human lupus inclusions and interferon.

Raji cells, a human B lymphoblastoid cell line of Burkitt lymphoma origin, formed lupus inclusions when grown in a medium conditioned by the growth of Raji cells whose DNA thymidine residues had been unifilarly (single-strandedly) substituted with bromodeoxyuridine. Ultracentrifugation of this medium in excess of that required to remove Epstein-Barr virus and all other known mammalian viruses did not prevent the formation of the inclusions, and treatment of the conditioned medium with pronase destroyed the activity. These results demonstrate the presence of a protein that is secreted from bromodeoxyuridine-substituted Raji cells and is capable of inducing nonbromodeoxyuridine-substituted cells to form lupus inclusions. Interferon (100 units per milliliter) was found in the conditioned medium. Inclusions also formed in Raji cells grown in fresh medium supplemented with human leukocyte or fibroblast interferon (100 units per milliliter).

Bromodeoxyuridine↗
PubMed1979

An analysis of electrophoretic and microcolumn methods for the separation of hemoglobins A and A2.

Parameters of three techniques for quantitating hemoglobin A2 were studied in order to identify problems affecting repeatability and then to compare intertechnique results under selected conditions. Satisfactory repeatability with cellulose acetate electrophoresis and scanning densitometry required an applicator that delivers a constant volume of sample. For cellulose acetate electrophoresis/elution and chromatographic assays a sophisticated absorption spectrophotometer and pH meter are necessary. Even with the most carefully chosen conditions significant intertechnique variation occurs. Although the colums are the most repeatable, trailing (a problem usually associated with hemoglobin electrophoresis) has also been demonstrated with column chromatography. Isoelectric focusing demonstrated the copresence of hemoglobin A and hemoglobin A2 in all trail fractions between the two major peaks and in some fractions of each peak. Standards of low protein concentration could be prepared from column fractions identified by isoelectric focusing as containing only hemoglobin A or hemoglobin A2. Such standards would be useful for assessing the accuracy of hemoglobin A2 quantitation.

Chromatography, Ion Exchange↗
PubMed1978

Population screening for beta-thalassemia minor. Report of cooperative trials based on two approaches.

In a cooperative intrastate program based upon experience with sickle-cell anemia screening, the authors explored the feasibility of applying hemoglobin electrophoresis for detection of beta-thalassemia gene carriers. Initially, blood samples collected in capillary tubes were analyzed by cellulose acetate electrophoresis with densitometric quantitation of hemoglobin A2 (Hb A2), followed by selective spectrophotometric quantitation. This approach proved insufficiently specific or reproducible. Follow-up hematologic and family studies of presumptive beta-thalassemia gene carriers indicated that coordinate measurement of erythrocytic indices and Hb A2 values would have discriminated a subpopulation with a high incidence of beta-thalassemia trait more specifically. This approach was tested prospectively by the use of 731 venous blood samples collected in a county with a large population of Mediterranean ancestry. Of 31 individuals (4.2%) with presumptive thalassemia trait, 13 returned for a repeat testing, and the initial results for 11 were confirmed. These findings lend support to an empirical screening sequence suggested by Pearson (erythrocytic indices followed by Hb A2 quantitation), but they also indicate that a significant subpopulation of beta-thalassemia gene carriers with limited phenotypic expression may elude detection in any single-pass approach.

Blood Protein Electrophoresis↗