Stanford T. Roodman

Chronic Hepatitis B in Asymptomatic Homosexual Men with Antibody to the Human Immunodeficiency Virus

Excerpt The discovery of the human immunodeficiency virus (formerly known as HTLV-III/LAV) as the causative agent of the acquired immunodeficiency syndrome (AIDS) has led to the commercial availabi...

Thymosin-α1 stimulates maturation of CD34+ stem cells into CD3+4+ cells in an in vitro thymic epithelia organ coculture model

The effect of thymosin-alpha1 on thymopoiesis is largely unknown. Thymosin is found in the cortical and medullary thymic epithelia, as well as in nurse cells; thus, it is hypothesized that thymosin may affect both early and late stage of thymocyte maturation. In this study, the effect of thymosin-alpha1 on thymopoiesis was determined by coculturing in vitro CD34+ stem cells (SC) with allogeneic cultured thymic epithelia fragments (CTEF) for 1-4 weeks and analyzing T-cell maturation by flow cytometry. Thymosin-alpha1 significantly enhanced the cell number (e.g., proliferation) of mononuclear cells obtained at 2 and 4 weeks of the SC-CTEF cocultures (P < 0.01 and < 0.05, respectively). In particular, thymosin-alpha1 stimulated expression of CD3+ cells at 3 and 4 weeks (P < 0.05). The predominant subpopulation increased by thymosin stimulation was single positive mature CD4+ cells, which was confirmed to occur within the SC-CTEF thymic organ tissue by laser confocal immunofluorescence microscopy. Thymosin stimulation tended to enhance IL-7 synthesis, critical cytokine in the maturation of thymocytes. In summary, this is the first study to demonstrate that thymosin-alpha1 enhanced thymopoiesis of CD34+ stem cells in humans using an in vitro model of differentiation using stem cells and cultured thymic epithelial fragments cocultures. Furthermore, the thymosin significantly increased expression of CD3+4+ T cells.

A Medical Follow-Up of the Health Effects of Long-Term Exposure to 2,3,7,8-Tetrachlorodibenzo-p-Dioxin

The human health effects of long-term exposure to low levels of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) have not been well established. The results of a prior study showed that persons exposed to TCDD had depressed cell-mediated immunity, and 18 of 51 persons had anergy or relative anergy on skin testing. This paper presents the results of a medical follow-up on participants who were reported to be anergic or relatively anergic in the earlier study. None of the participants in the follow-up study was anergic, and only one exposed and one unexposed participant were relatively anergic. Several technical and biological possibilities for the difference in results of the two studies are presented. The possibility that recovery from the effects of TCDD exposure caused the differing results is the least plausible explanation for the changes in the skin test results.

Immunoglobulin, complement, and immune complex levels during a migraine attack.

SYNOPSIS

Inhibition of Thymopoiesis of CD34+ Cell Maturation by HIV‐1 in an In Vitro CD34+ Cell and Thymic Epithelial Organ Culture Model

The mechanisms by which HIV‐1 affects thymopoiesis were determined by preincubating CD34+ cells or cultured thymic epithelial (CTE) cells with lymphotropic (T‐) and monotropic (M‐) strains of HIV‐1 in an in vitro CTE organ and CD34+ cell coculture model that allows for analysis of development of thymocytes and mature T cells.

Isolation and purification of human C1q from plasma

C1q was purified to homogeneity from human plasma by a 3-step purification procedure. Plasma was euglobulin precipitated, and the redissolved precipitate chromatographed on a rabbit IgG-Sepharose column. The 1 M NaCl buffer eluate was passed directly through a rabbit anti-human IgG-Sepharose affinity column. C1q freed of IgG was present in the flow through. The rationale for this scheme to remove IgG free and that bound to C1q is discussed. Overall recovery of C1q was about 40% with IgG less than 4 microgram/mg C1q. In SDS-polyacrylamide electrophoresis with non-reducing conditions bands at 52,000 and 42,000 daltons were demonstrated while with reducing conditions bands at 26,000, 24,000 and 20,000 daltons were found as reported by others. C1q was found to be stable at 4 degrees C for 1 year in a 1 M NaCl, 0.4 M Tris, 10% sucrose, 0.005 M EDTA, 0.02% NaN3, pH 8.6 buffer.

T Cell Differentiation/Maturation of CD34+ Stem Cells from HIV-Seropositive Hemophiliacs in Cultured Thymic Epithelial Fragments

The clinical manifestations of AIDS are predominantly due to the cellular and humoral immune dysfunction caused by HIV infection, and thymic dysplasia caused by HIV infection probably contributes to the T cell lymphopenia. In the present study, T cell differentiation and/or maturation was assessed when enriched CD34+ stem cells (SCs or SC) purified from bone marrow of HIV‐seropositive hemophiliacs were cocultured with allogeneic cultured thymic epithelial fragments (CTEFs).

Characterization of RNP and Sm ribonucleoprotein nuclear antigens

Antibodies to the RNase-sensitive RNP and to the RNase-resistant Sm nuclear antigens were used to affinity purify these antigens from a saline extract of rabbit thymus acetone powder. Determination of the protein subunits recovered by either glycine-HCl, pH 2.8, or 2.5 M MgCl2 elution on gradient sodium dodecyl sulfate-polyacrylamide electrophoresis containing mercaptoethanol revealed that RNP was composed of five proteins with mol. wts from 10,000 to 15,000 whereas Sm contained the same or similar five chains plus six additional subunits with mol. wts from 21,000 to 42,000. RNase treatment of the thymus extract increased the recovery in Sm of the same bands compared to untreated extract. Thus, RNP and Sm appear to have different numbers of protein components and RNP may be a subset of Sm. Sucrose gradient centrifugation of the 125I-labeled, pH 2.8 eluted antigens gave peaks of 3 and 6S for RNP and Sm, respectively. Sucrose gradient centrifugation of the crude untreated thymus extract followed by quantitative single radial immunodiffusion analysis of each fraction produced a broad peak from 16S to the top of the gradient while pretreatment of the extract with RNase resulted in a discrete 6S peak. These results indicate that in rabbit thymus acetone powder native RNP and Sm exist as larger polydisperse complexes with additional material including RNA and that after acid elution or RNase treatment the antigens are found in a smaller monodisperse form.

Detection of immune complexes using a solid-phase C1q polystyrene ball assay☆

A polystyrene ball C1q solid-phase assay (PSB C1q SPA) has been developed for quantitating immune complexes in human serum. While similar to the previously reported solid-phase C1q assays in principle, the use of polystyrene balls with a specular finish has resulted in an assay with significantly improved accuracy, sensitivity and reproducibility. The sensitivity of the assay based on the amount of AHGG bound per microgram C1q added was approximately 12-fold higher in the PSB assay compared to the polystyrene tube solid-phase C1q method. In order to correct for variable background contributions in different samples, values obtained with heat-inactivated C1q were subtracted from each experimental result. Reproducibility studies yielded a coefficient of variation (CV) of 10 and 4% in day-to-day assays using 25 microgram and 100 microgram AHGG/ml normal serum, respectively compared to 11--15% for the tube technique. Within run measurements gave a CV of 37 and 20% at the low and high levels of AHGG. Aggregated human gamma-globulin (AHGG) was used as a model immune complex and when chromatographed on Biogel A-15m yielded major fractions at greater than or equal to 15,000,000 and 150,000 daltons. Maximum binding of AHGG to PSB C1q occurred with aggregates greater than 15,000,000 daltons. Optimum binding of human albumin-anti-albumin complexes in the PSB C1q SPA occurred at a molar ratio of 1 : 1.5. The size distribution of this complex active in the assay determined by sucrose density gradients was 14--32 S with peaks at 21 and 27 S. A normal range of immune complexes was determined as 15 +/- 8 microgram AHGG equivalents (+/- 2 S.D., n = 65). Approximately 70% of rheumatoid arthritis, linear scleroderma, vasculitis, Sjögrens and glomerulonephritis and 40% of SLE patients were above 2 S.D. of normal. SLE patients demonstrated elevations in immune complexes only during periods of increased disease severity. The assay was a useful monitor of plasmapheresis in an SLE patient, showing decreases in immune complexes after each plasmapheresis. DNA did not interfere with AHGG binding whereas lipemia prevented detection of added AHGG.

Temperature-sensitive binding of solid phase C1q to aggregated human immunoglobulin G

The first component of complement (C1q) coupled to Sepharose by cyanogen bromide was found not to bind aggregated human gamma-globulin or immune complexes at room temperature, whereas at 4 degrees C binding was nearly complete. The temperature sensitivity of solid phase C1q binding was reversible. Elution of aggregated human gamma-globulin bound at 42 degrees C was possible by raising the temperature to 23 degrees C. However, free C1q or C1q adsorbed onto polystyrene balls could bind immune complex-like material at both 23 and 4 degrees C. The conformational restraints of C1q covalently coupled to a solid support may not allow functional activity at elevated temperatures.